Anti-CD68 antibody(AB125212)

Immunohistochemistry of microglial marker in mouse brain

Cortical staining for the microglial marker, CD68 increased as a function of age in rTg4510 animals, but remained unchanged in tTA animals. Scale bar, 200 μm, inset scale bar, 50 μm.

(After Figure 10 A of Wes et al.)

Wes, P.D. PLoS One. 2014 Aug 25;9(8):e106050. doi: 10.1371/journal.pone.0106050. eCollection 2014 Reproduced under the Creative Commons license http://creativecommons.org/licenses/by/4.0/

False colour image of Western blot: Anti-CD68 antibody staining at 0.2 μg/ml, shown in green; Mouse anti-Alpha Tubulin [DM1A] (ab7291) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab125212 was shown to bind specifically to CD68. A band was observed at 95-102 kDa in wild-type RAW 264.7 cell lysates with no signal observed at this size in CD68 knockout cell line ab280047 (knockout cell lysate ab280106). To generate this image, wild-type and CD68 knockout RAW 264.7 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution before incubation with primary antibodies overnight at 4°C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) at 1/20000 dilution.

All lanes: Western blot - Anti-CD68 antibody (ab125212) at 0.2 µg/mL

Lane 1: Wild-type RAW 264.7 cell lysate at 20 µg

Lane 2: CD68 knockout RAW 264.7 cell lysate at 20 µg

Lane 3: Mouse spleen cell lysate at 20 µg

Lane 4: Neuro-2a cell lysate at 20 µg

Performed under reducing conditions.

Predicted band size: 37 kDa

Observed band size: 95-102 kDa

Immunohistochemical analysis of paraffin-embedded Rat lung tissue labeling CD68 with ab125212 at 2 ?g/ml. Heat mediated antigen retrieval was performed in EDTA buffer (pH 8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with ab125212 overnight at 4°C. Peroxidase Conjugated Goat Anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using a HRP conjugated Rabbit IgG with DAB as the chromogen.