Objective
To test 3 new patients sera (Katia Adsvar, JoseM Gutierrez, Pablito Macatangay).
Materials
- NF155-transfected HEK cells, grown in coverslips and freezed
- CNTN1-transfected HEK cells, grown in coverslips and freezed
- Sera from 2 positive controls (LAC and HVH1) and3 new patients
Procedure
- Take one coverslip per condition (8)
- Prepare sera 1:40 in goat serum. Add a drop (40µl) of each preparation to the parafilm. Incubate for 1h
- Wash 3 times with PBS
- Incubate primary antibody (commercial): (1) mouse-anti-CNTN1 1:500 + (2) Chicken-anti-neurofascin155 1:2000 (dilution maked in the last ICC)
- Wash 3 times with PBS
- Prepare secondary antibody: (A) GAH IgG-594 1:500 + GAC-488 1:500 + (B) GAH IgG-594 1:500 + GAM-488 1:500. Add a drop and incubate 1h.
- Wash 3 times with H2O
- Mount with Vectashield (with DAPI)
Results & conclusions
Patients sera is negative for both antigens. Antibody against CNTN is not working in this ICC, we’ll do some tests.
