20140221 CNTN1 and NF155 ICC

Objective

To test a new patient

Materials

• NF155-transfected HEK cells, grown in coverslips the day before
• CNTN1-transfected HEK cells, grown in coverslips the day before
• Sera from 2 positive controls (LAC and HVH1) and new patient JSC

Procedure

1. Take one coverslip per condition (4)
2. Prepare sera 1:40 in goat serum. Add a drop (40µl) of each preparation to the parafilm. Incubate for 1h
3. Wash 3 times with PBS
4. Incubate primary antibody (commercial): (1) mouse-anti-CNTN1 1:500 + (2) Chicken-anti-neurofascin155 1:2000
5. Wash 3 times with PBS
6. 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.
7. Wash 3 times with H2O
8. Mount with Vectashield (with DAPI)

Results & conclusions

The patient is negative for both antibodies (CNTN1+NF155).