High sensitivity streptavidin biotin detection kits for mouse primary antibodies. 8 ml Kits including chromogen. Ready-to-use blocking solution, sec. antibody, enzym conjugate.
Assay & Detection
Paraffin-embedded tissue sections are first deparaffinised and rehydrated. Background staining caused by unspecific binding of the primary or secondary antibody 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 biotinylated secondary antibody is applied and incubated. This secondary antibody functions as a link between primary antibody and the streptavidinalkaline phosphatase-conjugate (“Streptavidin-AP-Conjugate“). A second washing is followed by the application of this conjugate. It binds to the biotin at the secondary antibody. Any excess of unbound streptavidin-AP-conjugate 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 Fast Red (included only in kit KDB-10020) leads to a magenta-red product of reaction at the place of the target antigen. Other suitable chromogens are Permanent AP Red (magenta-red), New Fuchsin (magenta-red) or NBT (blue-black) with its substrate BCIP. Staining procedure 1. Blocking Solution (protein block, Reagent 1) (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 2 min. 5. Biotinylated Secondary Antibody, Mouse (Reagent 2, yellow) 10-15 min. 6. Washing with wash buffer 3 x 2 min. 7. Streptavidin-AP-Conjugate (Reagent 3, red) 10-15 min. 8. Washing with wash buffer 3 x 2 min. 9. Fast Red, Permanent AP Red, NBT/BCIP or New Fuchsin 10-20 min. (Controlling the colour intensity via light microscope is recommended.) 10. Stopping the reaction with distilled H2O when the desired colour intensity is attained 11. Counterstaining and blueing 12. Mounting: aqueous with Fast Red, permanent with Permanent AP Red, NBT/BCIP or New Fuchsin
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
1 Kit (500ml / 5000 Tests)
2 - 8°C
High sensitivity broad spectrum streptavidin biotin detection kits for mouse primary antibodies. 8 ml kits including chromogen. Ready-to-use blocking solution, sec. antibody, enzyme conjugate.
Product Page Updated
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.