Detection of the G-CSFR protein in Western blot analyses. 1 × 10⁷ neutrophils were lysed. The G-CSFRs were immunoprecipitated with MoAb 129, separated by SDS-PAGE using an 8% wt/vol polyacrylamide gel, and blotted onto nitrocellulose membranes. Detection was performed with anti–G-CSFR antibody (MoAb 129; IgG1) and HRP-conjugated goat-antimouse IgG. The immunoblot was developed by the enhanced chemiluminescence method following the manufacturer's guidelines (ECL; Amersham). Lanes 1 and 2, patients with severe congenital neutropenia (patients with point mutations in the cytoplasmic domain of the G-CSFR mRNA). Lane 3, Healthy control. The arrows point out the G-CSFR proteins.