Missense mutations in LRRK2 (
leucine-rich repeat
kinase 2) are a major cause of PD (
Parkinson's disease). Several
antibodies against LRRK2 have been developed, but results using these polyclonal
antibodies have varied widely leading to conflicting conclusions. To address this challenge, the Michael J. Fox Foundation for Parkinson's Research generated a number of
monoclonal antibodies targeting
epitopes across the
LRRK2 protein. In the present paper, we report optimized protocols and results for ten
monoclonal antibodies for immunoblotting, immunohistochemistry, immunoprecipitation and
kinase activity assays, in rat, mouse and human brain tissue. Several efficacious
antibodies were identified, but results demonstrate that the mouse monoclonal N241A/34 is suitable for most applications, with the best overall rabbit
monoclonal antibody being c41-2. These
antibodies produced a dominant band of the expected size via immunoblotting and a lack of labelling in tissue derived from LRRK2-knockout animals under optimized conditions. A significant proportion of
LRRK2 protein localizes to insoluble fractions and no evidence of truncated
LRRK2 protein was detected in any fraction from rodent or human tissues. An assay was developed for the robust detection of LRRK2
kinase activity directly from frozen mouse and human brain tissue, but precipitous declines in activity were observed that corresponded to increasing post-mortem intervals and processing times. Finally, we demonstrate the highest levels of brain-localized LRRK2 in the striatum, but note differential expression patterns between rat and mouse in both striatum and cortex. Anti-LRRK2
monoclonal antibodies that are unlimited in availability together with the proposed standardized protocols should aid in the definition of LRRK2 function in both health and disease.