Application of glutaraldehyde to in-cell Western assay for normalization

Normalization is essential to the in-cell Western (ICW) assay, a near-infrared immunocytoblot for protein analysis. Here we report that cells reacted with glutaraldehyde fluoresced in the near-infrared region of the spectrum, and the intensity of fluorescence was directly proportional to cell number over a range from 3125 to 100,000 cells per well. We took advantage of this property to develop a method for quantification of cells, and applied it to the ICW assay for normalization. The application of glutaraldehyde may make the ICW assay more popular due to the reduced cost and simplified procedure.