Product Introduction:

This kit is suitable for immunohistochemical test of mouse IgG source as primary antibody. FITC fluorescence detection. SABC is designed for immunohistochemical and other immune assays to show the distribution of antigens in tissues and cells. StreptAvidin (StreptAvidin) is a protein extracted from Streptomyces, like avidin, has a very high affinity for biotin, and avidin is an alkaline protein (PI=10) that can be modified to become a neutral protein. The isoelectric point of streptavidin is close to neutral, and the non-specific adsorption to tissues and cells is very low. The background of streptavidin based immunohistochemical method is very low. SABC, or StreptAvidin - Biotin Complex, can form a complex of about 100 FITCs and about 50 streptavidins. A large number of FITCs will guarantee high sensitivity of SABC.

Operation steps: (Take the hot repair of paraffin sections as an example, please centrifuge the trace reagent and use it after)

1. Routine dewaxing of slices to water.

2. Optional steps:

A. Heat repair antigen, dip the slice into 0.01M sodium citrate buffer (PH6.0), heat in electric oven or microwave oven to boil, then power off, interval 5-10 minutes, repeat 1-2 times. After cooling, wash 1-2 times with PBS (pH7.2-7.4). b, enzyme digestion, drop digestive solution, 37℃ for 10 minutes, PBS (pH7.2-7.4) washing 2-3 times. c. Skip this step and go to the next step.

3. Add 5%BSA sealer and leave at room temperature for 20 minutes. Discard excess liquid and leave to wash.

4. Dilute the primary antibody with diluent in a certain proportion (the diluted primary antibody can be stored at 4℃ for one week), or you can purchase the antibody diluent separately. Drop diluted primary antibody and incubate at 37 ℃ for about 1 hour or 20℃ for 2 hours, or 4℃ overnight. PBS (pH7.2-7.4) was washed 3 times for 2 minutes each time (the dilution, incubation time and temperature of the primary antibody were directly related to the staining intensity and background. Generally speaking, when the positive staining intensity is not enough, the concentration of primary antibody can be increased and the incubation time can be prolonged. When the background is too high, it can reduce the concentration of primary antibody and shorten the incubation time).

5. According to the amount of use, dilute the Bio-Rabbit anti-goat IgG concentrate by 1:100 into the working liquid (1ml dilute with 10ul Bio-rabbit anti-goat IgG concentrate, mix well to form the working liquid. This working liquid can be stored at 4℃ for one week). Drop Bio-Rabbit anti-goat IgG working liquid,20-37℃ for 30 minutes. Wash with PBS (pH7.2-7.4) for 3 times, 2 minutes each time.

6. According to the amount of use, the SBC-FITC concentrate is diluted with diluent at 1:100 to the working liquid (1ml diluent is added to 10ul SBC-FITC concentrate and mixed to form the SBC-FITC working liquid, which can be stored at 4℃ for one week). Add the SBC-FITC working solution and incubate at 20-37℃ for 30 minutes (away from light). Wash with PBS (pH7.2-7.4) 4 times for 5 minutes each time.

7. Add anti-fluorescence attenuation sealant to seal the film. Fluorescence microscope observation. For cell crawling tablets, after fixation, rinsed twice with PBS, incubated with 0.5% Trxiton X-100 at room temperature for 20 minutes, rinsed twice with PBS, followed by step 3 above. For frozen slices, after fixation, rinse twice with PBS, then follow step 2 above.

Note:

If the background is too high, after the SABC reaction, the slices are washed with PBST (pH7.2-7.4) added with 0.01-0.02% TWEEN-20 for 4 times, PBS for 2 times, and then the slices are sealed for observation.

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