43 Year-Old Human Lung NKX2.1, AGER, and ACTA2 Confocal Imaging

Gene
Nkx2-1 LungGens Ensemble Genecards NCBI The Human Protein Atlas
AGER LungGens Ensemble Genecards NCBI The Human Protein Atlas
Acta2 LungGens Ensemble Genecards NCBI The Human Protein Atlas

Donor Tissue Kindly Provided by Dr. Gloria Pryhuber from the University of Rochester Medical Center

Pryhuber_Lab

Immunofluorescence-  NKX2.1, AGER, and ACTA2

 Purpose:  Stain slides of 7µm frozen sections of 43-year-old human lung, courtesy of Dr. Scott Randell, University of North Carolina, for NKX2.1, AGER, and ACTA2 with antigen retrieval.

Day 1

  1. For Frozen tissue, rinse 2X in PBS, then 5 min in 4% PFA/PBS, then rinse 1X in PBS.
  2. Briefly equilibrate slides in Antigen Retrieval Buffer.
  3. Antigen retrieval, pH 6.0 (times will vary according to microwave).
  4. 10 mM sodium citrate, pH 6.0, and heat in a microwave at 96oC. *We usually do a series of three runs (6-7 minutes each run) to equal the time/temp because of evaporation (refill coplin jars with dH2O).
  5. Microwave according to instructions on microwave.
  6. Cool on countertop, 15 min.
  7. Rinse with dH2O
  8. 1X PBS, 5 min.
  9. Block in 4% Donkey serum/PBS-T, 2 hours at RT.
  10. For Rabbit anti- NKX2.1  (RB TTF-1 1231, Lot# 0912A) dilute 1:1000, For rat anti-