Fluorescent Staining of Cryosections
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| Materials |
|---|
| PBS pH 7.4 |
| Blocking buffer (1% BSA, 1% serum and 0.1% Triton X-100 in
PBS) |
| Corresponding 1st and 2nd antibodies |
| Barrier pen |
| Mounting medium |
Procedure:
- Thaw the slides at room temperature for 10-20 min
- The slides are washed for 10 min in PBS
- Block non-specific staining by incubating samples in blocking buffer for 30 minutes at RT
- Surround the tissue with a hydrophobic barrier using a barrier pen
- Apply primary antibodies diluted in blocking buffer according to manufacturer’s instructions overnight at 4° C (now all incubations are carried out using a humidified chamber)
- The next day slides are washed in PBS 3 times for 15 min
- The slides are incubated in the corresponding 2ary antibody solution in blocking buffer for 1h at RT
- Finally the slides are washed three times in PBS and mounted