The Fast AP One-Step Polymer anti-Mouse/Rabbit is designed for the qualitative detection of antigens in fixed paraffin-embedded tissue sections, in frozen tissue sections, and in cytological samples. It was developed for use in combination with mono- and polyclonal primary antibodies and sera obtained from mice or rabbit. The kit can be used for examining tissues fixed in different solutions, e.g. formalin (neutrally buffered), B5, Bouin, ethanol, or HOPE. It is intended for in vitro diagnostic use.
The purpose of the immunohistochemical staining is to make tissue and cell antigens visible. The Fast AP One-Step Polymer anti-Mouse/Rabbit is a highly sensitive detection reagent intended for use in immunohistochemistry and immunocytochemistry. The enzyme polymer consists of several molecules of secondary antibodies covalently bound to several molecules of alkaline phosphatase (AP). Visualisation occurs via an enzymesubstrate reaction in the presence of a colorising reagent which permits microscopical analysis. The test system is suitable for the detection of mono- and polyclonal primary antibodies and sera obtained from mice or rabbit. Cross-reactivity with primary antibodies from rat has been observed. In contrast to other detection techniques, which often use the streptavidin-biotin system the Fast AP One-Step Polymer anti-Mouse/Rabbit avoids the problem of background staining caused by endogenous biotin in the tissue.
Principle of method:
Paraffin-embedded tissue sections are first deparaffinised and rehydrated. Background staining caused by unspecific binding of the primary antibody or the secondary antibody in the AP OneStep Polymer is minimized by incubation with a protein blocking solution. This step can be omitted if the primary antibodies are diluted in an appropriate buffer. The next step is incubation with the specific primary antibody. After washing, the AP One-Step Polymer is applied and incubated. Any excess of unbound polymer is thoroughly washed away after incubation. The addition of the chromogenic substrate starts the enzymatic reaction of the alkaline phosphatase which leads to colour precipitation where the primary antibody is bound. The colour can be observed with a light microscope. The chromogen used determines the colour. The chromogen Permanent AP Red leads to the formation of a magentared product of reaction at the place of the target antigen.
Reagents provided:
100 ml Fast AP One-Step Polymer anti-Mouse/Rabbit (ready-to-use) Substrate systems recommended: Permanaent AP Red kit Materials required but not supplied Positive and negative control tissue Xylene or suitable substitutes Ethanol, distilled H2O Reagents for enzyme digestion or heat pre-treatment Wash buffer PBS or TBS Blocking Solution (for protein blocking, optional) Pink PAP Pen Primary antibody (user-defined) Primary antibody diluent Negative control reagent Chromogenic substrate Counter stain solution Mounting medium Cover slips
Storage and handling:
The solution should be stored at 2-8°C without further dilution. Please store the reagent in a dark place and do not freeze it. Under these conditions the solution is stable up to the expiry date. It should not be used after the expiry date. A positive and a negative control have to be carried out in parallel to the test material. If you observe unusual staining or other deviations from the expected results which could possibly be caused by the reagent, please contact our technical support
Reagent preparation:
Reagents should be at room temperature when used. Deparaffinise and rehydrate paraffin-embedded tissue sections. Pre-treatment (optional) with HIER (Heat Induced Epitope Retrieval) or enzymatic digestion. Tissue sections have to be completely covered with the different reagents in order to avoid drying out.
Procedure:
1. Blocking Solution (This step is optional.) 5 min. 2. Washing with wash buffer 1 x 2 min. 3. Primary antibody (optimally diluted) or negative control reagent 30-60 min. 4. Washing with wash buffer 3 x 5 min. 5. AP One-Step Polymer anti-Mouse/Rabbit 30 Min. 6. Washing with wash buffer 3 x 2 min. 7. Permanent AP Red, 10-20 min. (Controlling the colour intensity via light microscope is recommended.) 8. Stopping the reaction with distilled H2O when the desired colour intensity is attained 9. Counterstaining and blueing 10. Mounting: permanent or aqueous with Permanent AP Red Kit
Expected results:
During the reaction of the substrate with alkaline phosphatase in the presence of a chromogen, a coloured precipitate is formed at the location of the bound primary antibody. This reaction only takes place if the target antigen is existent in the tissue. The chromogen used determines the colour of the precipitate. The analysis is carried out using a light microscope.
Trouble shooting:
If you observe unusual staining or other deviations from the expected results which could possibly be caused by the reagents, please read these instructions carefully, contact our technical support. No staining on an actually positive control slide: 1. Reagents were not used in the proper order. 2. Chromogenic substrate solution was too old. 3. Bleaching because chromogen and mounting medium are incompatible. 4. The antigen/epitope in the tissue was insufficiently accessible to the primary antibody. Try a pre-treatment such as heat pre-treatment or enzyme digestion. If you used a pre-treatment it should be extended. 5. Primary antibody not from mouse or rabbit but from a different species. 6. The antigen/epitope was not stable in the fixation and/or pre-treatment procedure used. Try another fixation or pre-treatment. Weak staining: 1. Inadequate fixation or overfixation. 2. Incomplete deparaffinisation. 3. The antigen/epitope in the tissue was insufficiently accessible to the primary antibody. If you used heat pre-treatment or enzyme digestion it should be extended. 4. Excessive incubation with Blocking Solution or insufficient washing after this step. 5. Too much wash buffer remains on the slides after washing, diluting the reagents applied in the next step. 6. Incubation times were too short or primary antibody concentration too low. 7. Chromogenic substrate solution was too old. Nonspecific background staining or overstaining: 1. Incomplete deparaffinisation. 2. Excessive tissue adhesive on slides. 3. Insufficient washing especially after the incubation with the enzyme polymer or the chromogenic substrate solution. These washings are critical. 4. Tissue was allowed to (partially) dry out with reagents on. 5. Unspecific binding of the primary antibody. Please use Blocking Solution or dilute the primary antibody in appropriate diluents. 6. Incubation time of the primary antibody was too long or primary antibody concentration too high. 7. Incubation time of the chromogenic substrate solution was too long or reaction temperature too high (e.g. if temperature in the laboratory is high).
Quality Control:
We recommend carrying out a positive and a negative control with every staining run. The positive control permits the validation of appropriate processing of the sample. If the negative control has a positive result, this points to unspecific staining.
Performance characteristics:
Studies have been conducted to evaluate the performance of the kit reagents. The product has been found to be suitable for the intended use
Limitations of procedure:
Immunohistochemistry is a complex method in which histological as well as immunological detection methods are combined. Tissue processing and handling prior to immunostaining, for example variations in fixation and embedding or the inherent nature of the tissue can cause inconsistent results (Nadji and Morales, 1983). Inadequate counterstaining and mounting can influence the interpretation of the results. The colour intensity of the reaction product can decrease with time, especially when exposed to light. Sanbio guarantees that the product will meet all requirements described from its shipping date until its expiry date, as long as the product is correctly stored and utilized. No additional guarantees can be given. Under no circumstances shall Sanbio be liable for any damages arising out of the use of the reagent provided.
Precautions:
Use by qualified personnel only. Wear protective clothing to avoid eye, skin or mucous membrane contact with the reagents. In case of a reagent coming into contact with a sensitive area, wash the area with large amounts of water. Microbial contamination of the reagents must be avoided, since otherwise non-specific staining might appear. ProClin 300 is used for stabilisation. A Material safety data sheet (MSDS) is available upon request.
AEC Single Solution is a ready-to-use solution intended for immunohistochemical and in situ-hybridisation staining procedures with horse radish peroxidase (HRP). AEC (3-Amino-9-ethylcarbazol) leads to the formation of a red-brown precipitate at the location of the target antigen or target nucleic acid. The precipitate is insoluble in aqueous mounting media and can be observed by light microscopy. AEC Single Solution is especially useful when a high sensitivity is desired.
100 ml AEC Single Solution (ready-to-use) 2 Dropper Bottles
Storage and handling:
The solution should be stored at 2-8°C without furt her dilution. Please store the reagent in a dark place and do not freeze it. Under these conditions the solution is stable up to the expiry date indicated on the label. Do not use product after the expiry date. AEC Single Solution is a ready-to-use solution. Preparation of a working solution as in other chromogenic substrates is not necessary. A positive and a negative control have to be carried out in parallel to the test material. If you observe unusual staining or other deviations from the expected results which could possibly be caused by the kit reagents please contact our technical support.
Reagent preparation:
The solution is ready-to-use. AEC Single Solution can be used directly from the refrigerator and should be stored again at 2-8°C aft er use. When using the small package (8 ml) please directly drop from the bottle. When using the 100 ml package (MON-APP209) please transfer up to 8 ml of the AEC Single Solution into one of the provided dropper bottles. The transferred solution is stable for many weeks if stored at 2-8°C. The volume required for several staining runs should be transferred so that the 100 ml stock bottle has to be opened only a few times. If you would like to pipette the solution use a clean vial from which you pipette. Remaining quantities should not be filled back into the bottle but disposed as hazardous material.
Procedure:
1) Rinse the slide with wash buffer after the previous incubation step. 2) Apply the AEC Single Solution to the slide. Incubate for 3-6 minutes. (Incubation time can be extended up to 30 minutes, if desired.) 3) Rinse with distilled H2O. 4) Counterstain with haematoxylin for about 30 seconds up to 5 minutes (depending on the desired staining intensity). 5) Rinse with distilled H2O. 6) Blueing in tap water for at least 5 minutes. 7) Mount with an aqueous mounting medium.
Expected results:
During the reaction of the substrate with horse radish peroxidase in presence of the chromogen AEC, a red-brown precipitate is formed at the location of the target antigen or nucleic acid. The precipitate is insoluble in aqueous solvents and can be observed by light microscopy.
Trouble shooting:
If you observe unusual staining or other deviations from the expected results please read these instructions carefully, contact our technical support. Also refer to the instructions of the detection systems for guidance on general troubleshooting.
Quality Control:
We recommend carrying out a positive and a negative control with every staining run. The positive control permits the validation of appropriate processing of the sample. If the negative control has a positive result, this points to unspecific staining. Please refer to the instructions of the detection system for guidance on general quality control procedures.
Performance characteristics:
Studies have been conducted to evaluate the performance of the kit reagents. The product has been found to be suitable for the intended use
Limitations of procedure:
Immunohistochemistry is a complex method in which histological as well as immunological detection methods are combined. Tissue processing and handling prior to immunostaining, for example variations in fixation and embedding or the inherent nature of the tissue can cause inconsistent results (Nadji and Morales, 1983). In some tissues endogenous peroxidase activity may cause non-specific staining. The enzyme activity should be blocked by incubation with hydrogen peroxide solution (H2O2 solution). The step is carried out before incubation with primary antibody but after dewaxing and rehydration. Background staining due to endogenous biotin can be blocked through an avidin-biotin blocking step prior to the primary antibody incubation step. The coloured precipitate formed by AEC is soluble in organic solvents. The tissue sections therefore have to be counterstained with aqueous solutions (e. g. Gills or Mayers haematoxylin) and mounted with aqueous mounting media. The colour intensity of the reaction product can decrease with time, especially when exposed to light. The staining reaction itself can be influenced in the same way when carried out in strong light. Sanbio guarantees that the product will meet all requirements described from its shipping date until its expiry date, as long as the product is correctly stored and utilized. No additional guarantees can be given. Under no circumstances shall Sanbio be liable for any damages arising out of the use of the reagent provided.
Precautions:
Use by qualified personnel only. AEC (3-Amino-9-ethylcarbazol) and the solvents used are considered hazardous materials. Material safety data sheets (MSDS) are available upon request. Wear protective clothing to avoid contact of reagent or specimen with eye, skin or mucous membrane. In case of reagent or specimen coming into contact with a sensitive area, wash the area with large amounts of water. Oxidising substances, e. g. metals, dust, bacteria or glass devices can influence the stability of AEC Single Solution. Such contaminations have to be avoided. Non-consumed solution needs to be discarded as dangerous substance.
Permanent HRP Green Kit is intended for immunohistochemical and in situ-hybridisation staining procedures with horse radish peroxidase (HRP). It results in the formation of a green precipitate at the location of the target antigen or target nucleic acid. The precipitate is insoluble in organic solvents and can be observed by light microscopy. The HRP Green precipitate shows a very good contrast to red chromogenic substrates used with alkaline phosphatase detection systems and is therefore especially recommended for double stains. Permanent HRP Green Kit is intended for research use only.
100 ml HRP Green Substrate Buffer 3 ml HRP Green Chromogen 1 Dilution Vial
Storage and handling:
The solutions should be stored at 2-8°C without fur ther dilution. Please store the reagents in a dark place and do not freeze them. Under these conditions the solutions are stable up to the expiry date indicated on the label. Do not use product after the expiry date. The working solution should be prepared freshly at the day of use. It is stable for at least 4 hours. Excess working solution should be disposed. A positive and a negative control have to be carried out in parallel to the test material. If you observe unusual staining or other deviations from the expected results which could possibly be caused by the kit reagents please contact our technical support.
Reagent preparation:
1) Pipette 1 ml HRP Green Substrate Buffer into the provided dilution vial. 2) Add 1 drop (30 µl) of HRP Green Chromogen. Mix thoroughly. 3) The resulting working solution is stable for at least 4 hours. If you want to prepare other quantities of the working solution, please use the same ratio HRP Green Substrate and HRP Chromogen.
Procedure:
1) Rinse the slide with wash buffer after the previous incubation step. 2) Apply the Permanent HRP Green working solution onto the slide. Incubate for 2-10 minutes. 3) Rinse with distilled H2O. 4) Counterstain with haematoxylin for about 30 seconds up to 5 minutes (depending on the desired staining intensity). 5) Rinse with distilled H2O. 6) Blueing in tap water for 5 minutes. 7) Dehydrate through a graded series of ethanol and clear in xylene. Do not exceed incubation times of 30 sec per dehydration step. Use only high purity xylene. Mount with a permanent mounting medium. Note: The colour intensity can be intensified by increasing the chromogen concentration (up to 3 drops or 90 µl chromogen) in the working solution. Lithium carbonate may have a negative effect on the staining result. We recommend to only bluing in tap water. Occasionally precipitates may appear in the HRP Green Chromogen solution. This doesnt affect the staining result.
Expected results:
During the reaction of the substrate with horse radish peroxidase in presence of the Permanent HRP Green chromogen, a green precipitate is formed at the location of the target antigen or nucleic acid. The precipitate is insoluble in organic solvents and can be observed by light microscopy.
Trouble shooting:
If you observe unusual staining or other deviations from the expected results please read these instructions carefully, contact our technical support. Also refer to the instructions of the detection systems for guidance on general troubleshooting.
Quality Control:
We recommend carrying out a positive and a negative control with every staining run. The positive control permits the validation of appropriate processing of the sample. If the negative control has a positive result, this points to unspecific staining. Please refer to the instructions of the detection system for guidance on general quality control procedures.
Performance characteristics:
Studies have been conducted to evaluate the performance of the kit reagents. The product has been found to be suitable for the intended use
Limitations of procedure:
Too long incubation steps at the final dehydration serious can diminish the staining intensity. Also low grade xylene and some forms of recycled alcohol can have a negative effect on the staining result. In double stain procedures we recommend to use Permanent HRP Green as the last chromogen. Immunohistochemistry is a complex method in which histological as well as immunological detection methods are combined. Tissue processing and handling prior to immunostaining, for example variations in fixation and embedding or the inherent nature of the tissue can cause inconsistent results (Nadji and Morales, 1983). Sanbio guarantees that the product will meet all requirements described from its shipping date until its expiry date, as long as the product is correctly stored and utilized. No additional guarantees can be given. Under no circumstances shall Sanbio be liable for any damages arising out of the use of the reagent provided.
Precautions:
Use by qualified personnel only. Wear protective clothing to avoid contact of reagents or specimen with eye, skin or mucous membrane. In case of a reagent or specimen coming into contact with a sensitive area, wash the area with large amounts of water. Microbial contamination of the reagents must be avoided, since otherwise non-specific staining may occur. Material safety data sheets (MSDS) are available upon request.
Paraformaldehyde (PFA) is the polymerization product of formaldehyde with a typical degree of polymerization of 8100 units. Paraformaldehyde is not a fixative itself; it must be depolymerized to formaldehyde in solution. In cell culture, a typical formaldehyde fixing procedure would involve using a 4% formaldehyde solution in phosphate buffered saline (PBS) on ice for 10 minutes. Fixing ensures that sample cell structures stay intact and that antigens are immobilized, while ideally still permitting unfettered access of antibodies to target antigens. CAS 30525-89-4, soluble in water, pH 7.0-7.6 at 25°C. The solution should be clear, colorless, with no precipitate.
Principle of method:
Fixing cells for immunohistochemistry (IHC)
Reagents provided:
500 ml 4 % Paraformaldehyde (PFA) solution in PBS
Storage and handling:
Store at -20°C for one year
Trouble shooting:
Adjust pH to 7.2-7.6 with NaH2PO4 if pH
Precautions:
Causes serious eye damage. Suspected of causing cancer. May cause an allergic skin reaction. Causes skin irritation. Avoid breathing dust/fume/gas/mist/vapors/spray. Wear protective gloves/protective clothing/eye protection/face protection. Use personal protective equipment as required. IF IN EYES: Rinse cautiously with water for several minutes. Remove contact lenses, if present and easy to do. Continue rinsing. Store locked up. Dispose of contents/container in accordance with local/regional/national/international regulations
CryoCompound freezing medium is an aqueous based frozen tissue embedding medium designed to support tissue blocks in cryostat sectioning. Formulated to promote rapid freezing, enhanced sectioning and consistent results at a working temperature of -20° C. The ready-to-use Cryocompound is available in different colors: MON-APP900C (Transparant/colorless), MON-APP900B (Blue), MON-APP900Y (Yellow), MON-APP900O (Orange), MON-APP900G (green), MON-APP900R (Red) or Multicolorkit MON-APP900BYGR (Blue, Yellow, Green, Red).
Principle of method:
Freezing medium
Reagents provided:
100 ml Cryocompound, freezing medium (ready-to-use)
Storage and handling:
Store at room temperature
Procedure:
1. Place few drops of CryoCompound (depends on the size of the sample to be embedded) onto the center of the bottom of cryomold. Be careful to select the proper size embedding mold according to the size of the sample to be embedded. 2. Try to avoid the formation of air bubbles. Remove any bubbles inside the CryoCopound. This is important because the air bubbles will create problems when cutting sections. Air bubbles create freeze- thaw-freeze cycle and ice crystal will form inside of it and result in a very bad morphology due to the ice crystal artefact. 3. Let it settle for 15-30 seconds to allow the CryoCompound to completely wet the surface of the tissue. Hardening of the CryoCompound included the sample (it will happen in 0.5-1 minute) before preparing the CryoCompoundsample-block.
CryoCompound freezing medium is an aqueous based frozen tissue embedding medium designed to support tissue blocks in cryostat sectioning. Formulated to promote rapid freezing, enhanced sectioning and consistent results at a working temperature of -20° C. The ready-to-use Cryocompound is available in different colors: MON-APP900C (Transparant/colorless), MON-APP900B (Blue), MON-APP900Y (Yellow), MON-APP900O (Orange), MON-APP900G (green), MON-APP900R (Red) or Multicolorkit MON-APP900BYGR (Blue, Yellow, Green, Red).
Principle of method:
Freezing medium
Reagents provided:
4x 100 ml Cryocompound, freezing medium (ready-to-use)
Storage and handling:
Store at room temperature
Procedure:
1. Place few drops of CryoCompound (depends on the size of the sample to be embedded) onto the center of the bottom of cryomold. Be careful to select the proper size embedding mold according to the size of the sample to be embedded. 2. Try to avoid the formation of air bubbles. Remove any bubbles inside the CryoCopound. This is important because the air bubbles will create problems when cutting sections. Air bubbles create freeze- thaw-freeze cycle and ice crystal will form inside of it and result in a very bad morphology due to the ice crystal artefact. 3. Let it settle for 15-30 seconds to allow the CryoCompound to completely wet the surface of the tissue. Hardening of the CryoCompound included the sample (it will happen in 0.5-1 minute) before preparing the CryoCompoundsample-block.
CryoCompound freezing medium is an aqueous based frozen tissue embedding medium designed to support tissue blocks in cryostat sectioning. Formulated to promote rapid freezing, enhanced sectioning and consistent results at a working temperature of -20° C. The ready-to-use Cryocompound is available in different colors: MON-APP900C (Transparant/colorless), MON-APP900B (Blue), MON-APP900Y (Yellow), MON-APP900O (Orange), MON-APP900G (green), MON-APP900R (Red) or Multicolorkit MON-APP900BYGR (Blue, Yellow, Green, Red).
Principle of method:
Freezing medium
Reagents provided:
100 ml Cryocompound, freezing medium (ready-to-use)
Storage and handling:
Store at room temperature
Procedure:
1. Place few drops of CryoCompound (depends on the size of the sample to be embedded) onto the center of the bottom of cryomold. Be careful to select the proper size embedding mold according to the size of the sample to be embedded. 2. Try to avoid the formation of air bubbles. Remove any bubbles inside the CryoCopound. This is important because the air bubbles will create problems when cutting sections. Air bubbles create freeze- thaw-freeze cycle and ice crystal will form inside of it and result in a very bad morphology due to the ice crystal artefact. 3. Let it settle for 15-30 seconds to allow the CryoCompound to completely wet the surface of the tissue. Hardening of the CryoCompound included the sample (it will happen in 0.5-1 minute) before preparing the CryoCompoundsample-block.
CryoCompound freezing medium is an aqueous based frozen tissue embedding medium designed to support tissue blocks in cryostat sectioning. Formulated to promote rapid freezing, enhanced sectioning and consistent results at a working temperature of -20° C. The ready-to-use Cryocompound is available in different colors: MON-APP900C (Transparant/colorless), MON-APP900B (Blue), MON-APP900Y (Yellow), MON-APP900O (Orange), MON-APP900G (green), MON-APP900R (Red) or Multicolorkit MON-APP900BYGR (Blue, Yellow, Green, Red).
Principle of method:
Freezing medium
Reagents provided:
100 ml Cryocompound, freezing medium (ready-to-use)
Storage and handling:
Store at room temperature
Procedure:
1. Place few drops of CryoCompound (depends on the size of the sample to be embedded) onto the center of the bottom of cryomold. Be careful to select the proper size embedding mold according to the size of the sample to be embedded. 2. Try to avoid the formation of air bubbles. Remove any bubbles inside the CryoCopound. This is important because the air bubbles will create problems when cutting sections. Air bubbles create freeze- thaw-freeze cycle and ice crystal will form inside of it and result in a very bad morphology due to the ice crystal artefact. 3. Let it settle for 15-30 seconds to allow the CryoCompound to completely wet the surface of the tissue. Hardening of the CryoCompound included the sample (it will happen in 0.5-1 minute) before preparing the CryoCompoundsample-block.
CryoCompound freezing medium is an aqueous based frozen tissue embedding medium designed to support tissue blocks in cryostat sectioning. Formulated to promote rapid freezing, enhanced sectioning and consistent results at a working temperature of -20° C. The ready-to-use Cryocompound is available in different colors: MON-APP900C (Transparant/colorless), MON-APP900B (Blue), MON-APP900Y (Yellow), MON-APP900O (Orange), MON-APP900G (green), MON-APP900R (Red) or Multicolorkit MON-APP900BYGR (Blue, Yellow, Green, Red).
Principle of method:
Freezing medium
Reagents provided:
100 ml Cryocompound, freezing medium (ready-to-use)
Storage and handling:
Store at room temperature
Procedure:
1. Place few drops of CryoCompound (depends on the size of the sample to be embedded) onto the center of the bottom of cryomold. Be careful to select the proper size embedding mold according to the size of the sample to be embedded. 2. Try to avoid the formation of air bubbles. Remove any bubbles inside the CryoCopound. This is important because the air bubbles will create problems when cutting sections. Air bubbles create freeze- thaw-freeze cycle and ice crystal will form inside of it and result in a very bad morphology due to the ice crystal artefact. 3. Let it settle for 15-30 seconds to allow the CryoCompound to completely wet the surface of the tissue. Hardening of the CryoCompound included the sample (it will happen in 0.5-1 minute) before preparing the CryoCompoundsample-block.
CryoCompound freezing medium is an aqueous based frozen tissue embedding medium designed to support tissue blocks in cryostat sectioning. Formulated to promote rapid freezing, enhanced sectioning and consistent results at a working temperature of -20° C. The ready-to-use Cryocompound is available in different colors: MON-APP900C (Transparant/colorless), MON-APP900B (Blue), MON-APP900Y (Yellow), MON-APP900O (Orange), MON-APP900G (green), MON-APP900R (Red) or Multicolorkit MON-APP900BYGR (Blue, Yellow, Green, Red).
Principle of method:
Freezing medium
Reagents provided:
100 ml Cryocompound, freezing medium (ready-to-use)
Storage and handling:
Store at room temperature
Procedure:
1. Place few drops of CryoCompound (depends on the size of the sample to be embedded) onto the center of the bottom of cryomold. Be careful to select the proper size embedding mold according to the size of the sample to be embedded. 2. Try to avoid the formation of air bubbles. Remove any bubbles inside the CryoCopound. This is important because the air bubbles will create problems when cutting sections. Air bubbles create freeze- thaw-freeze cycle and ice crystal will form inside of it and result in a very bad morphology due to the ice crystal artefact. 3. Let it settle for 15-30 seconds to allow the CryoCompound to completely wet the surface of the tissue. Hardening of the CryoCompound included the sample (it will happen in 0.5-1 minute) before preparing the CryoCompoundsample-block.
CryoCompound freezing medium is an aqueous based frozen tissue embedding medium designed to support tissue blocks in cryostat sectioning. Formulated to promote rapid freezing, enhanced sectioning and consistent results at a working temperature of -20° C. The ready-to-use Cryocompound is available in different colors: MON-APP900C (Transparant/colorless), MON-APP900B (Blue), MON-APP900Y (Yellow), MON-APP900O (Orange), MON-APP900G (green), MON-APP900R (Red) or Multicolorkit MON-APP900BYGR (Blue, Yellow, Green, Red).
Principle of method:
Freezing medium
Reagents provided:
100 ml Cryocompound, freezing medium (ready-to-use)
Storage and handling:
Store at room temperature
Procedure:
1. Place few drops of CryoCompound (depends on the size of the sample to be embedded) onto the center of the bottom of cryomold. Be careful to select the proper size embedding mold according to the size of the sample to be embedded. 2. Try to avoid the formation of air bubbles. Remove any bubbles inside the CryoCopound. This is important because the air bubbles will create problems when cutting sections. Air bubbles create freeze- thaw-freeze cycle and ice crystal will form inside of it and result in a very bad morphology due to the ice crystal artefact. 3. Let it settle for 15-30 seconds to allow the CryoCompound to completely wet the surface of the tissue. Hardening of the CryoCompound included the sample (it will happen in 0.5-1 minute) before preparing the CryoCompoundsample-block.
The aqua mounting medium is a widely used for coverslipping from aqueous solutions and is non-fluorescing and has an antifade component to increase the viewing time of the specimen. Use Aqua-Poly/Mount with most fluorescent dyes and stains including DAB, Alkaline Phosphatase, Fast Red, AEC (aminoethylcarbazole) and a variety of other chromogens to enhance and retain fluorescent intensity. The mounting medium (water based) can be used for frozen sections, fat stains, chromogens for immunohistochemistry and in situ hybridization as well as other applications requiring a water-soluble mounting medium.
Principle of method:
Coverslipping from aqueous solutions
Reagents provided:
50 ml Mounting medium (water based), ready-to-use.
Storage and handling:
Store at room temperature
Procedure:
1. Prepare slides as required. 2. Prior to coverslipping rinse the slides in distilled or deionized water. 3. Place the bottle upside down with the end cap attached in a container before use. This will help clear of tiny bubbles. 4. Blot excess water from the slide without drying the tissue specimen. The tissue must be moist prior to mounting. 5. Apply enough drops of aqua mounting medium on the tissue section so that the tissue is completely covered. 6. Carefully lower the coverslip at an angle while gently applying pressure to force any excess medium and air bubbles away from the tissue and out from under the coverslip. 7. Slides can be dried at room temperature or at 4°C or in an oven but the temperature hight is depending what type of staining is used. 8. Allow the aqua mounting medium to dry before examine under the microscope.
For in-vitro Diagnostic Use. The BrightVision two components detection system peroxidase Goat Anti-Mouse/Rabbit IgG HRP, is intended for use in immunohistochemistry for the detection of mouse or rabbit antibodies.
The BrightVision detection system peroxidase Goat Anti- Mouse/Rabbit HRP, is a Ready-to-Use system that has been manufactured to give an optimal staining, when using the protocol advised in this IFU. Prior to staining some routine fixed, paraffin-embedding tissue sections should be subjected to pre-treatment (HIER or digestive enzyme). The BrightVision detection system detects Mouse or Rabbit bound to an antigen in tissue sections. This polymer-complex is then visualized with a suitable substrate/chromogen (not provided).
Principle of method:
Two steps detection system goat anti-mouse/rabbit HRP
Reagents provided:
Post-blocking (ready-to-use) 55 ml and Polymer Goat Anti-Mouse/Rabbit HRP (ready-to-use) 55 ml.
Storage and handling:
2-8°C and in the dark
Procedure:
1. Deparaffinize and rehydrate tissue section (slide/tissue peparing), 2. Wash Aqua dest (Wash; 2x 5 min), 3. If applicable, HIER or digestive enzyme (pre-treatment), 4. Wash buffer (PBS or TBS buffer; 2x 5 min), 5. H2O2 (conc3%) (Tissue preparing; 10 min), 6. Wash buffer (PBS or TBS buffer; 2x 5 min), 7. Primary mouse or rabbit antibody (Antibody; 30 min), 8. Wash buffer (PBS or TBS buffer; 2x 5 min), 9. Detection system, step 1, post-blocking (Post-blocking; 15 min), 10. Wash buffer (PBS or TBS buffer; 2x 5 min), 11. Detection system, step 2, polymer Mouse/Rabbit HRP (Labeled polymer; 30 min), 12. Wash buffer (PBS or TBS buffer; 2x 5 min), 13. If applicable Substrate (DAB), 14. Wash aqua dest (Wash; 2x 2 min), 15. Counterstain, dehydrate and coverslip (Auxiliary)
Quality Control:
A positive control, negative control and reagent control are needed and processed in the same way as the unknow specimen slide to interpret staining results.
For in-vitro Diagnostic Use. The BrightVision two components detection system peroxidase Goat Anti-Mouse/Rabbit IgG HRP, is intended for use in immunohistochemistry for the detection of mouse or rabbit antibodies.
The BrightVision detection system peroxidase Goat Anti- Mouse/Rabbit HRP, is a Ready-to-Use system that has been manufactured to give an optimal staining, when using the protocol advised in this IFU. Prior to staining some routine fixed, paraffin-embedding tissue sections should be subjected to pre-treatment (HIER or digestive enzyme). The BrightVision detection system detects Mouse or Rabbit bound to an antigen in tissue sections. This polymer-complex is then visualized with a suitable substrate/chromogen (not provided).
Principle of method:
Two steps detection system goat anti-mouse/rabbit HRP
Reagents provided:
Post-blocking (ready-to-use) (Gold) 55 ml and Polymer Goat Anti-Mouse/Rabbit HRP (ready-to-use) (Ruby) 55 ml.
Storage and handling:
2-8°C and in the dark
Procedure:
1. Deparaffinize and rehydrate tissue section (slide/tissue peparing), 2. Wash Aqua dest (Wash; 2x 5 min), 3. If applicable, HIER or digestive enzyme (pre-treatment), 4. Wash buffer (PBS or TBS buffer; 2x 5 min), 5. H2O2 (conc3%) (Tissue preparing; 10 min), 6. Wash buffer (PBS or TBS buffer; 2x 5 min), 7. Primary mouse or rabbit antibody (Antibody; 30 min), 8. Wash buffer (PBS or TBS buffer; 2x 5 min), 9. Detection system, step 1, post-blocking (Post-blocking; 15 min), 10. Wash buffer (PBS or TBS buffer; 2x 5 min), 11. Detection system, step 2, polymer Mouse/Rabbit HRP (Labeled polymer; 30 min), 12. Wash buffer (PBS or TBS buffer; 2x 5 min), 13. If applicable Substrate (DAB), 14. Wash aqua dest (Wash; 2x 2 min), 15. Counterstain, dehydrate and coverslip (Auxiliary)
Quality Control:
A positive control, negative control and reagent control are needed and processed in the same way as the unknow specimen slide to interpret staining results.
For in-vitro Diagnostic Use. The BrightVision two components detection system peroxidase Goat Anti-Mouse/Rabbit IgG AP, is intended for use in immunohistochemistry for the detection of mouse or rabbit antibodies.
The BrightVision detection system peroxidase Goat Anti- Mouse/Rabbit AP, is a Ready-to-Use system that has been manufactured to give an optimal staining, when using the protocol advised in this IFU. Prior to staining some routine fixed, paraffin-embedding tissue sections should be subjected to pre-treatment (HIER or digestive enzyme). The BrightVision detection system detects Mouse or Rabbit bound to an antigen in tissue sections. This polymer-complex is then visualized with a suitable substrate/chromogen (not provided).
Principle of method:
Two steps detection system goat anti-mouse/rabbit AP
Reagents provided:
Post-blocking (ready-to-use) 55 ml and Polymer Goat Anti-Mouse/Rabbit AP (ready-to-use) 55 ml.
Storage and handling:
2-8°C and in the dark
Procedure:
1. Deparaffinize and rehydrate tissue section (slide/tissue peparing), 2. Wash Aqua dest (Wash; 2x 5 min), 3. If applicable, HIER or digestive enzyme (pre-treatment), 4. Wash buffer (PBS or TBS buffer; 2x 5 min), 5. H2O2 (conc3%) (Tissue preparing; 10 min), 6. Wash buffer (PBS or TBS buffer; 2x 5 min), 7. Primary mouse or rabbit antibody (Antibody; 30 min), 8. Wash buffer (PBS or TBS buffer; 2x 5 min), 9. Detection system, step 1, post-blocking (Post-blocking; 15 min), 10. Wash buffer (PBS or TBS buffer; 2x 5 min), 11. Detection system, step 2, polymer Mouse/Rabbit AP (Labeled polymer; 30 min), 12. Wash buffer (PBS or TBS buffer; 2x 5 min), 13. If applicable Substrate (DAB), 14. Wash aqua dest (Wash; 2x 2 min), 15. Counterstain, dehydrate and coverslip (Auxiliary)
Quality Control:
A positive control, negative control and reagent control are needed and processed in the same way as the unknow specimen slide to interpret staining results.
For in-vitro Diagnostic Use. The BrightVision two components detection system peroxidase Goat Anti-Mouse/Rabbit IgG AP, is intended for use in immunohistochemistry for the detection of mouse or rabbit antibodies.
The BrightVision detection system peroxidase Goat Anti- Mouse/Rabbit AP, is a Ready-to-Use system that has been manufactured to give an optimal staining, when using the protocol advised in this IFU. Prior to staining some routine fixed, paraffin-embedding tissue sections should be subjected to pre-treatment (HIER or digestive enzyme). The BrightVision detection system detects Mouse or Rabbit bound to an antigen in tissue sections. This polymer-complex is then visualized with a suitable substrate/chromogen (not provided).
Principle of method:
Two steps detection system goat anti-mouse/rabbit AP
Reagents provided:
Post-blocking (ready-to-use) (Gold) 55 ml and Polymer Goat Anti-Mouse/Rabbit AP (ready-to-use) (Ruby) 55 ml.
Storage and handling:
2-8°C and in the dark
Procedure:
1. Deparaffinize and rehydrate tissue section (slide/tissue peparing), 2. Wash Aqua dest (Wash; 2x 5 min), 3. If applicable, HIER or digestive enzyme (pre-treatment), 4. Wash buffer (PBS or TBS buffer; 2x 5 min), 5. H2O2 (conc3%) (Tissue preparing; 10 min), 6. Wash buffer (PBS or TBS buffer; 2x 5 min), 7. Primary mouse or rabbit antibody (Antibody; 30 min), 8. Wash buffer (PBS or TBS buffer; 2x 5 min), 9. Detection system, step 1, post-blocking (Post-blocking; 15 min), 10. Wash buffer (PBS or TBS buffer; 2x 5 min), 11. Detection system, step 2, polymer Mouse/Rabbit AP (Labeled polymer; 30 min), 12. Wash buffer (PBS or TBS buffer; 2x 5 min), 13. If applicable Substrate (DAB), 14. Wash aqua dest (Wash; 2x 2 min), 15. Counterstain, dehydrate and coverslip (Auxiliary)
Quality Control:
A positive control, negative control and reagent control are needed and processed in the same way as the unknow specimen slide to interpret staining results.
For in-vitro Diagnostic Use. The BrightVision one step detection system peroxidase Goat Anti-Mouse/Rabbit IgG HRP, is intended for use in immunohistochemistry for the detection of mouse or rabbit antibodies.
The BrightVision detection system peroxidase Goat Anti- Mouse/Rabbit HRP, is a Ready-to-Use system that has been manufactured to give an optimal staining, when using the protocol advised in this IFU. Prior to staining some routine fixed, paraffin-embedding tissue sections should be subjected to pre-treatment (HIER or digestive enzyme). The BrightVision detection system detects Mouse or Rabbit bound to an antigen in tissue sections. This polymer-complex is then visualized with a suitable substrate/chromogen (not provided). Also available in 110 ml, 500 ml and 1000 ml.
Principle of method:
One step detection system peroxidase goat anti-mouse/rabbit IgG HRP
Reagents provided:
One step detection system Goat anti-Mouse/Rabbit HRP (Ready-to-use; 55 ml)
Storage and handling:
2-8°C and in the dark
Procedure:
1. Deparaffinize and rehydrate tissue section (slide/tissue peparing), 2. Wash Aqua dest (Wash; 2x 5 min), 3. If applicable, HIER or digestive enzyme (pre-treatment), 4. Wash buffer (PBS or TBS buffer; 2x 5 min), 5. H2O2 (conc3%) (Tissue preparing; 10 min), 6. Wash buffer (PBS or TBS buffer; 2x 5 min), 7. Primary mouse or rabbit antibody (Antibody; 30 min), 8. Wash buffer (PBS or TBS buffer; 2x 5 min), 9. Detection system, polymer HRP, (Labeled polymer; 30 min), 10. Wash buffer (PBS or TBS buffer; 2x 5 min), 11. Substrate (DAB; see applicable IFU), 12. Wash aqua dest (Wash; 2x 2 min), 13. Counterstain, dehydrate and coverslip (Auxiliary)
Quality Control:
A positive control, negative control and reagent control are needed and processed in the same way as the unknow specimen slide to interpret staining results.
For in-vitro Diagnostic Use. The BrightVision one step detection system Goat Anti-Mouse/Rabbit IgG AP, is intended for use in immunohistochemistry for the detection of mouse or rabbit antibodies.
The BrightVision detection system Goat Anti- Mouse/Rabbit AP, is a Ready-to-Use system that has been manufactured to give an optimal staining, when using the protocol advised in this IFU. Prior to staining some routine fixed, paraffin-embedding tissue sections should be subjected to pre-treatment (HIER or digestive enzyme). The BrightVision detection system detects Mouse or Rabbit bound to an antigen in tissue sections. This polymer-complex is then visualized with a suitable substrate/chromogen (not provided). Also available in 110 ml, 500 ml and 1000 ml.
Principle of method:
One step detection system goat anti-mouse/rabbit IgG AP
Reagents provided:
One step detection system Goat anti-Mouse/Rabbit AP (Ready-to-use; 55 ml)
Storage and handling:
2-8°C and in the dark
Procedure:
1. Deparaffinize and rehydrate tissue section (slide/tissue peparing), 2. Wash Aqua dest (Wash; 2x 5 min), 3. If applicable, HIER or digestive enzyme (pre-treatment), 4. Wash buffer (PBS or TBS buffer; 2x 5 min), 5. H2O2 (conc3%) (Tissue preparing; 10 min), 6. Wash buffer (PBS or TBS buffer; 2x 5 min), 7. Primary mouse or rabbit antibody (Antibody; 30 min), 8. Wash buffer (PBS or TBS buffer; 2x 5 min), 9. Detection system, polymer Mouse/Rabbit AP, (Labeled polymer; 30 min), 10. Wash buffer (TBS buffer; 2x 5 min), 11. Substrate (Fast Red / New Fuchsin, see applicable IFU), 12. Wash aqua dest (Wash; 2x 2 min), 13. Counterstain and coverslip with aqueous mounting medium.
Quality Control:
A positive control, negative control and reagent control are needed and processed in the same way as the unknow specimen slide to interpret staining results.
For in-vitro Diagnostic Use. The BrightVision one step detection system peroxidase Goat Anti-Mouse IgG HRP, is intended for use in immunohistochemistry for the detection of mouse antibodies.
The BrightVision detection system peroxidase Goat Anti- Mouse HRP, is a Ready-to-Use system that has been manufactured to give an optimal staining, when using the protocol advised in this IFU. Prior to staining some routine fixed, paraffin-embedding tissue sections should be subjected to pre-treatment (HIER or digestive enzyme). The BrightVision detection system detects Mouse bound to an antigen in tissue sections. This polymer-complex is then visualized with a suitable substrate/chromogen (not provided). Also available in 110 ml, 500 ml and 1000 ml.
Principle of method:
One step detection system peroxidase goat anti-mouse IgG HRP
Reagents provided:
One step detection system Goat anti-Mouse HRP (Ready-to-use; 55 ml)
Storage and handling:
2-8°C and in the dark
Procedure:
1. Deparaffinize and rehydrate tissue section (slide/tissue peparing), 2. Wash Aqua dest (Wash; 2x 5 min), 3. If applicable, HIER or digestive enzyme (pre-treatment), 4. Wash buffer (PBS or TBS buffer; 2x 5 min), 5. H2O2 (conc3%) (Tissue preparing; 10 min), 6. Wash buffer (PBS or TBS buffer; 2x 5 min), 7. Primary mouse antibody (Antibody; 30 min), 8. Wash buffer (PBS or TBS buffer; 2x 5 min), 9. Detection system, step 1, polymer Mouse, (Labeled polymer; 30 min), 10. Wash buffer (PBS or TBS buffer; 2x 5 min), 11. Substrate (DAB; see applicable IFU), 12. Wash aqua dest (Wash; 2x 2 min), 13. Counterstain, dehydrate and coverslip (Auxiliary)
Quality Control:
A positive control, negative control and reagent control are needed and processed in the same way as the unknow specimen slide to interpret staining results.
For in-vitro Diagnostic Use. The BrightVision one step detection system peroxidase Goat Anti-Mouse IgG AP, is intended for use in immunohistochemistry for the detection of mouse antibodies.
The BrightVision detection system Goat Anti- Mouse AP, is a Ready-to-Use system that has been manufactured to give an optimal staining, when using the protocol advised in this IFU. Prior to staining some routine fixed, paraffin-embedding tissue sections should be subjected to pre-treatment (HIER or digestive enzyme). The BrightVision detection system detects Mouse bound to an antigen in tissue sections. This polymer-complex is then visualized with a suitable substrate/chromogen (not provided). Also available in 110 ml, 500 ml and 1000 ml.
Principle of method:
One step detection system peroxidase goat anti-mouse IgG AP
Reagents provided:
One step detection system Goat anti-Mouse AP (Ready-to-use; 55 ml)
Storage and handling:
2-8°C and in the dark
Procedure:
1. Deparaffinize and rehydrate tissue section (slide/tissue peparing), 2. Wash Aqua dest (Wash; 2x 5 min), 3. If applicable, HIER or digestive enzyme (pre-treatment), 4. Wash buffer (PBS or TBS buffer; 2x 5 min), 5. H2O2 (conc3%) (Tissue preparing; 10 min), 6. Wash buffer (PBS or TBS buffer; 2x 5 min), 7. Primary mouse antibody (Antibody; 30 min), 8. Wash buffer (TBS buffer; 2x 5 min), 9. Detection system, polymer Mouse AP, (Labeled polymer; 30 min), 10. Wash buffer (TBS buffer; 2x 5 min), 11. Substrate (FAST Red / New Fuchsin; see applicable IFU), 12. Wash aqua dest (Wash; 2x 2 min), 13. Counterstain and coverslip with a aqueous mounting medium (Auxiliary)
Quality Control:
A positive control, negative control and reagent control are needed and processed in the same way as the unknow specimen slide to interpret staining results.
For in-vitro Diagnostic Use. The BrightVision one step detection system peroxidase Goat Anti-Rabbit IgG HRP, is intended for use in immunohistochemistry for the detection of rabbit antibodies.
The BrightVision detection system peroxidase Goat Anti-Rabbit HRP, is a Ready-to-Use system that has been manufactured to give an optimal staining, when using the protocol advised in this IFU. Prior to staining some routine fixed, paraffin-embedding tissue sections should be subjected to pre-treatment (HIER or digestive enzyme). The BrightVision detection system detects Rabbit bound to an antigen in tissue sections. This polymer-complex is then visualized with a suitable substrate/chromogen (not provided). Also available in 110 ml, 500 ml and 1000 ml.
Principle of method:
One step detection system peroxidase goat anti-rabbit IgG HRP
Reagents provided:
One step detection system Goat anti-Rabbit HRP (Ready-to-use; 55 ml)
Storage and handling:
2-8°C and in the dark
Procedure:
1. Deparaffinize and rehydrate tissue section (slide/tissue peparing), 2. Wash Aqua dest (Wash; 2x 5 min), 3. If applicable, HIER or digestive enzyme (pre-treatment), 4. H2O2 (conc3%) (Tissue preparing; 10 min), 5. Wash buffer (PBS or TBS buffer; 2x 5 min), 6. Primary rabbit antibody (Antibody; 30 min), 7. Wash buffer (PBS or TBS buffer; 2x 5 min), 8. Detection system, polymer Rabbit HRP, (Labeled polymer; 30 min), 9. Wash buffer (PBS or TBS buffer; 2x 5 min), 10. Substrate (DAB; see applicable IFU), 11. Wash aqua dest (Wash; 2x 2 min), 12. Counterstain, dehydrate and coverslip (Auxiliary)
Quality Control:
A positive control, negative control and reagent control are needed and processed in the same way as the unknow specimen slide to interpret staining results.
For in-vitro Diagnostic Use. The BrightVision one step detection system Goat Anti-Rabbit IgG AP, is intended for use in immunohistochemistry for the detection of rabbit antibodies.
The BrightVision detection system Goat Anti-Rabbit AP, is a Ready-to-Use system that has been manufactured to give an optimal staining, when using the protocol advised in this IFU. Prior to staining some routine fixed, paraffin-embedding tissue sections should be subjected to pre-treatment (HIER or digestive enzyme). The BrightVision detection system detects Rabbit bound to an antigen in tissue sections. This polymer-complex is then visualized with a suitable substrate/chromogen (not provided). Also available in 110 ml, 500 ml and 1000 ml.
Principle of method:
One step detection system goat anti-rabbit IgG AP
Reagents provided:
One step detection system Goat anti-Rabbit AP (Ready-to-use; 55 ml)
Storage and handling:
2-8°C and in the dark
Procedure:
1. Deparaffinize and rehydrate tissue section (slide/tissue peparing), 2. Wash Aqua dest (Wash; 2x 5 min), 3. If applicable, HIER or digestive enzyme (pre-treatment), 4. Wash buffer (PBS or TBS buffer; 2x 5 min), 5. H2O2 (conc3%) (Tissue preparing; 10 min), 6. Wash buffer (PBS or TBS buffer; 2x 5 min), 7. Primary rabbit antibody (Antibody; 30 min), 8. Wash buffer (TBS buffer; 2x 5 min), 9. Detection system, polymer Rabbit AP, (Labeled polymer; 30 min), 10. Wash buffer (TBS buffer; 2x 5 min), 11. Substrate (Fast Red / New Fuchsin; see applicable IFU), 12. Wash aqua dest (Wash; 2x 2 min), 12. Counterstain and coverslip with aqueous mounting medium.
Quality Control:
A positive control, negative control and reagent control are needed and processed in the same way as the unknow specimen slide to interpret staining results.
For in-vitro Diagnostic Use. The BrightVision Ultimate plus two steps component detection system peroxidase Goat Anti-Mouse/Rabbit IgG HRP and DAB, is intended for use in immunohistochemistry for the detection of mouse or rabbit antibodies.
The BrightVision Ultimate plus detection system, peroxidase Goat Anti-Mouse/Rabbit HRP plus DAB, is a Ready-to-Use system that has been manufactured to give an optimal staining, when using the protocol advised in this IFU. Prior to staining some routine fixed, paraffin-embedding tissue sections should be subjected to pre-treatment (HIER or digestive enzyme). The BrightVision Ultimate plus detection system detects Mouse or Rabbit bound to an antigen in tissue sections. This polymer-complex is then visualized with a suitable substrate/chromogen. The clinical interpretation of any staining or its absence should be determined by a qualified pathologist and complemented by morphologic studies; controls should be evaluated within the context of the patients clinical history and/or other diagnostic tests.
Principle of method:
BrightVision Ultimate plus, two steps detection system, Goat Anti-Mouse/Rabbit HRP (Ready-to-Use) plus DAB.
Reagents provided:
BrightVision Ultimate plus, two steps detection system, Goat Anti-Mouse/Rabbit HRP (Ready-to-Use) plus DAB 1. Post-blocking (ready-to-use): 55 ml 2. Polymer Goat Anti-Mouse/Rabbit HRP (Ready-to-Use): 55 ml 3. DAB Solution A: Buffered H2O2 (Ready-to-Use): 83 ml 4. DAB Solution B: Concentrated DAB solution: 4 ml
Storage and handling:
2-8°C and in the dark
Reagent preparation:
Preparation DAB: Add 40?l DAB Solution B (one drop) to 1 ml substrate Solution A, mix well. Volume and the quality of the Bright DAB has been formulized so they also can be used in automatic stainers, when a higher volume is required.
Procedure:
1. Deparaffinize and rehydrate tissue section (slide/tissue peparing), 2. Wash Aqua dest (Wash; 2x 5 min), 3. If applicable, HIER or digestive enzyme (pre-treatment), 4. Wash buffer (PBS or TBS buffer; 2x 5 min), 5. H2O2 (conc3%) (Tissue preparing; 10 min), 6. Wash buffer (PBS or TBS buffer; 2x 5 min), 7. Primary mouse or rabbit antibody (Antibody; 30 min), 8. Wash buffer (PBS or TBS buffer; 2x 5 min), 9. Detection system, step 1, post-blocking (Post-blocking; 15 min), 10 Wash buffer (PBS or TBS buffer; 2x 5 in). 11 Detection system, step 2, polymer Mouse/Rabbit HRP, (Labeled polymer; 30 min), 12. Wash buffer (PBS or TBS buffer; 2x 5 min), 13. Substrate (DAB; 8 min; Preparation DAB: Add 40?l DAB Solution B (one drop) to 1 ml substrate Solution A, mix well. Volume and the quality of the Bright DAB has been formulized so they also can be used in automatic stainers, when a higher volume is required.), 14. Wash aqua dest (Wash; 2x 2 min), 15. Counterstain, dyhydrate and coverslip (Auxiliary.
Quality Control:
A positive control, negative control and reagent control are needed and processed in the same way as the unknow specimen slide to interpret staining results.
For in-vitro Diagnostic Use. The BrightVision Ultimate one step component detection system peroxidase Goat Anti-Mouse/Rabbit IgG HRP and DAB, is intended for use in immunohistochemistry for the detection of mouse or rabbit antibodies.
The BrightVision Ultimate detection system, peroxidase Goat Anti-Mouse/Rabbit HRP plus DAB, is a Ready-to-Use system that has been manufactured to give an optimal staining, when using the protocol advised in this IFU. Prior to staining some routine fixed, paraffin-embedding tissue sections should be subjected to pre-treatment (HIER or digestive enzyme). The BrightVision Ultimate detection system detects Mouse or Rabbit bound to an antigen in tissue sections. This polymer-complex is then visualized with a suitable substrate/chromogen. The clinical interpretation of any staining or its absence should be determined by a qualified pathologist and complemented by morphologic studies; controls should be evaluated within the context of the patients clinical history and/or other diagnostic tests.
Principle of method:
BrightVision Ultimate, one step detection system, Goat Anti-Mouse/Rabbit HRP (Ready-to-Use) plus DAB.
Reagents provided:
BrightVision Ultimate plus, one detection system, Goat Anti-Mouse/Rabbit HRP (Ready-to-Use) plus DAB 1. one step detection, Goat Anti-Mouse/Rabbit HRP (Ready-to-Use): 55 ml 2. DAB Solution A: Buffered H2O2 (Ready-to-Use): 83 ml 4. DAB Solution B: Concentrated DAB solution: 4 ml
Storage and handling:
2-8°C and in the dark
Reagent preparation:
Preparation DAB: Add 40?l DAB Solution B (one drop) to 1 ml substrate Solution A, mix well. Volume and the quality of the Bright DAB has been formulized so they also can be used in automatic stainers, when a higher volume is required.
Procedure:
1. Deparaffinize and rehydrate tissue section (slide/tissue peparing), 2. Wash Aqua dest (Wash; 2x 5 min), 3. If applicable, HIER or digestive enzyme (pre-treatment), 4. Wash buffer (PBS or TBS buffer; 2x 5 min), 5. H2O2 (conc3%) (Tissue preparing; 10 min), 6. Wash buffer (PBS or TBS buffer; 2x 5 min), 7. Primary mouse or rabbit antibody (Antibody; 30 min), 8. Wash buffer (PBS or TBS buffer; 2x 5 min), 9 Detection system, polymer Mouse/Rabbit HRP, (Labeled polymer; 30 min), 10. Wash buffer (PBS or TBS buffer; 2x 5 min), 11. Substrate (DAB; 8 min.; Preparation DAB: Add 40?l DAB Solution B (one drop) to 1 ml substrate Solution A, mix well. Volume and the quality of the Bright DAB has been formulized so they also can be used in automatic stainers, when a higher volume is required.), 14. Wash aqua dest (Wash; 2x 2 min), 15. Counterstain, dyhydrate and coverslip (Auxiliary.
Quality Control:
A positive control, negative control and reagent control are needed and processed in the same way as the unknow specimen slide to interpret staining results.
For in-vitro Diagnostic Use. The BrightVision one step detection system Goat Anti-Mouse IgG HRP and Goat anti-rabbit IgG AP, is intended for use in immunohistochemistry for the detection of mouse or rabbit antibodies.
The BrightVision detection system, peroxidase Goat Anti-Mouse HRP/Goat anti-Rabbit AP, is a Ready-to-Use system that has been manufactured to give an optimal staining, when using the protocol advised in this IFU. Prior to staining some routine fixed, paraffin-embedding tissue sections should be subjected to pre-treatment (HIER or digestive enzyme). The BrightVision detection system detects Rabbit and Mouse bound to an antigen in tissue sections. This polymer-complex is then visualized with a suitable substrate/chromogen. The clinical interpretation of any staining or its absence should be determined by a qualified pathologist and complemented by morphologic studies; controls should be evaluated within the context of the patients clinical history and/or other diagnostic tests.
Principle of method:
BrightVision, one step detection system Goat Anti- Mouse HRP and Rabbit AP (Ready-to-Use).
Reagents provided:
BrightVision, One step detection system Goat anti-Mouse HRP / Goat anti-Rabbit AP (Ready-to-use; 55 ml)
Storage and handling:
2-8°C and in the dark
Procedure:
1. Deparaffinize and rehydrate tissue section (slide/tissue peparing), 2. Wash Aqua dest (Wash; 2x 5 min), 3. If applicable, HIER or digestive enzyme (pre-treatment), 4. Wash buffer (PBS or TBS buffer; 2x 5 min), 5. H2O2 (conc3%) (Tissue preparing; 10 min), 6. Wash buffer (PBS or TBS buffer; 2x 5 min), 7. Primary mouse and/or rabbit antibody (Antibody; 30 min), 8. Wash buffer (PBS or TBS buffer; 2x 5 min), 9 Detection system, polymer Mouse HRP/Rabbit AP, (Labeled polymer; 30 min), 10. Wash buffer (TBS buffer; 2x 5 min), 11. Substrate (DAB; see applicble IFU), 12. Wash aqua dest (Wash; 2x 2 min), 13. Substrate (Fast Red / New Fuchsin; see applicable IFU), 14. Wash aqua dest (Wash; 2x 2 min), 15. Counterstain and coverslip with aqueous mounting medium.
Quality Control:
A positive control, negative control and reagent control are needed and processed in the same way as the unknow specimen slide to interpret staining results.
Precautions:
Refer to SDS
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