A manual multiplex immunofluorescence method for investigating neurodegenerative diseases

The application of multiplex immunofluorescence to human post-mortem tissue would drive observational studies of selective vulnerability in neurodegenerative proteinopathies. Efficient elution of antibodies is critical for flexibility of antibody combinations and the ability to utilize a sample in multiple rounds of immunostaining. Here, we test two elution strategies for antibodies relevant to the study of selective vulnerability in neurodegenerative diseases in post-mortem human samples from both long-fixed and short-fixed tissue. Both 2-Mercaptoethanol/SDS-based and Urea/SDS/Glycine-based elution strategies work well with the antibodies selected, confirming observations from previous studies with other antibody types.