Objective: to test IgG reactivity against SH-SY5Y neuroblastoma derived neurons in patients iwht small fiber neuropathy
ICC protocol:
- Fix diferenciated SH-SY5Y cells with 4% PFA 5 min
- Wash 2 times with PBS 1x
- Block with goat serum 5% for 1h at RT
- Dilute serum 1/100 (IgG), 1/40 (IgM ) in blocking solution, incubate for 1h
- Wash with PBS 1x
- Commercial anti-NF Ab 1/200 diluted in blocking solution, incubate for 1h
- Wash with PBS 1x
- Secondary antibodies: GAH 488 IgM (1/500), GAC 594 (1/500)
- Wash with PBS 1x (not water)
- Vectashield mounting medium
Results: see table and photos
IgG:
| negative control | NEGATIVE |
|---|---|
| 1 | POSITIVE + (moderate somas, mild axons) |
| 2 | NEGATIVE |
| 33 | NEGATIVE |
| 45 | NEGATIVE |
| CNTN1+ control | POSITIVE ++ |
IgM:
| negative control | NEGATIVE |
|---|---|
| 2 | POSITIVE++ |
| 3 | NEGATIVE |
| 6 | NEGATIVE (only somas+) |
| 9 | NEGATIVE (only somas+) |
| 10 | NEGATIVE (only somas+) |
| 13 | NEGATIVE (only somas+) |
| 14 | NEGATIVE (only somas+) |
| 17 | NEGATIVE (only somas+) |
| 18 | NEGATIVE (only somas+) |
| 24 | NEGATIVE (only somas+) |
| 50 | NEGATIVE (only somas+) |
| 56 | NEGATIVE (only somas+) |
| control positive | POSITIVE |
