The Microglia Neurodegeneration Module Antibody Sampler Kit provides an economical means of detecting proteins identified as markers of microglial activity during neurodegeneration by western blot and/or immunofluorescence.
Specificity / Sensitivity
Each antibody in the Microglia Neurodegeneration Module Antibody Sampler Kit detects endogenous levels of its target protein. HS1 (D5A9) XP® Rabbit mAb (Rodent Specific) does not recognize human HS1 protein. HS1 has a calculated size of 54 kDa, but has an apparent molecular weight of 80 kDa on SDS-PAGE gels. Cathepsin B (D1C7Y) XP® Rabbit mAb recognizes endogenous levels of total cathepsin B protein and detects the heavy chain subunit of cathepsin B. Hydroxy-HIF-1α (Pro564) (D43B5) XP® Rabbit mAb detects endogenous levels of HIF-1α only when hydroxylated at Pro564 and may cross-react with other overexpressed proline hydroxylated proteins. Axl (C89E7) Rabbit mAb does not cross-react with Tyro3.
Source / Purification
Monoclonal antibodies are produced by immunizing animals with synthetic peptides corresponding to residues surrounding Leu310 of mouse HS1, Leu478 of human HIF-1α, the amino terminus of human Galectin-3/LGALS3, a hydroxypeptide surrounding Pro564 of human HIF-1α, and recombinant proteins specific to mouse ASC/TMS1, human Axl, human CD68, and the heavy chain subunit of human cathepsin B protein.
MultiMab™ rabbit monoclonal mix antibodies are prepared by combining individual rabbit monoclonal clones in optimized ratios for the approved applications. This product is optimized to detect CD68 as a monomer and a dimer by western blot and was produced by immunizing animals with recombinant human CD68 protein.
Distinct microglial activation states have been identified using RNA-seq data from a vast array of neurological disease and aging models. These activation states have been categorized into modules corresponding to proliferation, neurodegeneration, interferon-relation, LPS-relation, and many others (1). Previous work identifying markers of specific brain cell types using RNA-seq has shown HS1 and ASC/TMS1 to be useful and specific tools to study microglia (2). HS1 is a protein kinase substrate that is expressed only in tissues and cells of hematopoietic origin (3) and ASC/TMS1 has been found to be a critical component of inflammatory signaling where it associates with and activates caspase-1 in response to pro-inflammatory signals (4).
CD68 is a common marker for macrophage lineage cells; with expression found in the lysosome making it a useful marker for activated phagocytic microglia (5). Galectin-3 has been shown to regulate inflammatory response in neurodegenerative diseases, released by microglia in response to inflammatory stimuli (6). Cathepsin B is a widely expressed cysteine peptidase located in the lysosome as well as processed and secreted, playing a role in microglial-mediated neuronal death (7). Hypoxia inducible factor-1 (HIF-1α) is a transcription factor responsible for adaptation to low oxygen environments whose downstream effects have been shown in a number of neurodegenerative diseases. Under normoxic conditions, HIF-1α is proline hydroxylated leading to ubiquitin mediated degradation (8). Axl is a receptor tyrosine kinase that binds Gas6, stimulating regulatory effects on microglial phagocytic response to inflammatory stimuli (9).