IHC/ICC procedure for frozen sections, paraffin sections, cell smears and immunoblots.
1. Deparafinize and hydrate tissue sections through xylene or other clearing agents and graded alcohols.(For frozen sections or cell smears; use unfixed, acetone fixed or appropriate fixative for the antigen in question; for cell smears it may be necessary to permeabilize the cell by detergent, please refer to antibody protocol)
2. Wash 2-3 with distilled or deionized water.
3. Wash slide with Tris/saline buffer, followed by blocking with normal blocking serum, rinse with buffer.
4. Block with reagent S (streptavidin) for 5-10 minutes, rinse with buffer.
5. Now block with reagent B (Biotin) for 5-10 minutes, wash thoroughly with buffer.
Note: If antigen retrieval is required it can be applied after this stage.
6. Wash slide with PBS or Tris saline (with 0.02-0.05% nonionic detergent, Triton X100, Tween 20 or NP-40)
7. Follow instructions for IHC/ICC.
WB Procedure
1. After protein blocking step, soak blot in streptavidin reagent S, diluted 1:20 with tris/saline buffer for 5-10 minute.
2. Rinse blot with buffer; followed by soaking in diluted biotin reagent B (diluted 1:20) for 5-10 minutes.
3. Rinse with buffer.
These are guide lines, the optimum incubation times for these reagents and reactions should be determined by the individual lab.
PROPERTIES
Form
Liquid
Buffer
10mM PO₄, 200mM NaCl pH7.4, 10mg/ml BSA
Preservative
0.05% ProClin 300
Storage
Store at 2-8ºC.
Concentration
1 mg/ml (Please refer to the vial label for the specific concentration.)
Note
For laboratory research use only. Not for any clinical, therapeutic, or diagnostic use in humans or animals. Not for animal or human consumption.
Purchasers shall not, and agree not to enable third parties to, analyze, copy, reverse engineer or otherwise attempt to determine the structure or sequence of the product.
TARGET
Background
Some tissue sections contain endogenous biotin, biotin binding proteins, lections or non-specific binding substances. Tissues may also bind streptavidin, biotin, peroxidase biotin, peroxidase streptavidin in streptavidin biotin based Immunohistochemistry. These tissues will give high in the absence of biotinylated secondary antibodies. It may be necessary to block the tissue with streptavidin, followed by biotin. These reagents are added before the primary antibody.
Kit Components:
Ready-to-use Reagent S (Streptavidin) - 15ml
Ready-to-use Reagent B (Biotin) - 15ml
