The new lots of HRP-polymers contain a different red color which is more orange in order to distinguish HRP polymers and AP polymers. Properties and quality of the products will remain the same.
(HRP) Polymer Bulk Kit
High sensitivy polymer detection kits for mouse and rabbit primary antibodies. Ready-to-use Blocking Solution, PostBlock, Enzym Polymer.
Assay & Detection
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. Staining procedure 1. Peroxide blocking (3 % H2O2 solution) 10 min. 2. Washing with wash buffer 1 x 2 min. 3. Blocking Solution (protein block, Reagent 1) (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. PostBlock (Reagent 2, yellow) 20 min. 8. Washing with wash buffer 3 x 5 min. 9. HRP-polymer (Reagent 3, red) 30 min. 10. Washing with wash buffer 3 x 2 min. 11. AEC or DAB (Controlling the colour intensity via light microscope is recommended.) 5-15 min. 12. Stopping the reaction with distilled H2O when the desired colour intensity is attained 13. Counterstaining and blueing 14. 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 (“Blocking Solution”, provided with the kit). 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 enhancement reagent (“PostBlock“) is applied and incubated. A second washing is followed by the application of the HRP-polymer. 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.
Elias JM “Immunohistopathology – A practical Approach to Diagnosis” 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
1 Kit (100 ml /1000 Tests)
The solutions should be stored at 2-8°C without further 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. They should not be used after the expiry date.
The delivery time for this item is approximately 7-10 business days. If other deviations occur, this will be noted on the order confirmation or communicated via email. Please note, we do reserve the right to select the best packaging and shipping method in order to insure the stability of the product and allow efficient order tracking.