Assay & Detection

(HRP) One-Step Polymer anti-Mouse/Rabbit/Rat

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(HRP) One-Step Polymer anti-Mouse/Rabbit/Rat

Article No

KDB-10008

Application

IHC

Species Reactivity

mouse
rabbit
rat

Size

6 ml / 60 Tests

Product insert

Suggested protocols

Specifications

Application IHC
Article No KDB-10008
BioSite Brand BioSite Histo
Conjugation HRP
Description (HRP) One-Step Polymer anti-Mouse/Rabbit/Rat
Supplier BioSite Histo
Notes Enzym polymer single reagents with horse radish peroxidase. Ready-to-use
Product Type Assay & Detection
Protocol &S226; Reagents should be at room temperature when used. &S226; Deparaffinise and rehydrate paraffin-embedded tissue sections. &S226; Pre-treatment (optional) with HIER (Heat Induced Epitope Retrieval) or enzymatic digestion. &S226; Tissue sections have to be completely covered with the different reagents in order to avoid drying out Staining procedure 1. Peroxide blocking (3 % H2O2 solution) 10 min. 2. Washing with wash buffer 1 x 2 min. 3. Blocking Solution (This step is optional.) 5 min. 4. Washing with wash buffer 1 x 2 min. 5. Primary antibody (optimally diluted) or negative control reagent 30-60 min. 6. Washing with wash buffer 3 x 5 min. 7. HRP-polymer anti-Mouse/Rabbit 30 min. 8. Washing with wash buffer 3 x 2 min. 9. AEC or DAB (Controlling the colour intensity via light microscope is recommended.) 5-15 min. 10. Stopping the reaction with distilled H2O when the desired colour intensity is attained 11. Counterstaining and blueing 12. Mounting: aqueous with AEC, permanent with DAB Paraffin-embedded tissue sections are first deparaffinised and rehydrated. Endogenous peroxidase activity in the tissue may cause non-specific staining. This enzyme activity can be blocked by incubation with 3% H2O2-solution (peroxide block). Background staining caused by unspecific binding of the primary antibody or the secondary antibody in the HRP-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 HRP-polymer is applied and incubated. Any excess of unbound HRP-polymer is thoroughly washed away after incubation. The addition of the chromogenic substrate starts the enzymatic reaction of the peroxidase 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 AEC leads to the formation of a red-brown product of reaction at the place of the target antigen. The chromogen DAB forms a dark brown precipitate.
References Elias JM &S220;Immunohistopathology &S211; A practical Approach to Diagnosis&S221; ASCP Press 2003 Nadji M and Morales AR Ann N.Y. Acad Sci 420:134-9, 1983 Omata M et al. Am J Clin Pathol 73(5): 626-32, 1980
Size 6 ml / 60 Tests
Species Reactivity mouse, rabbit, rat
Storage The solutions 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 solutions are stable up to the expiry date. They should not be used after the expiry date.
Technical Specifications Single step polymer detection kit with horseradish peroxidase
Product Page Updated 2019-08-15T09:29:01.207Z

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