NeuroMab™ Anti-ABCA1 BBB Shuttle Antibody, Clone NDF4C2

Figure 1 shows a Western blot of activated RAW264.7 cells using antibodies against ABCA1.

RAW 264.7 cells were activated by incubation for 18 h with 1 μmol/L of TO901317. Cells were then lysed in RIPA buffer and proteins were separated on a 7.5% SDS-polyacrylamide gel followed by immunoblotting and staining with antibody NDF 4C2.

Figure 2 shows that antibodies can detect changes in ABCA1 abundance.

RAW 264.7 mouse macrophages were activated with the LXR activator TO901317. Equal amounts of cellular proteins from activated and non-activated cells were analyzed by western blot using antibody NDF4C2 followed by densitometry. A seven-fold increase in the abundance of ABCA1 was detected in RAW 264.7 cells.

Figure 3. Shows Western blot of activated or non-activated RAW 264.7 cells (A) or THP-1, HEK 293/hABCA1, 3T3, and RAW 264.7 cells (B) using antibodies against ABCA1.

Figure 4. Confocal microscopy of THP-1 cells stained with anti-ABCA1 antibody is shown.

Figure 5A. Graph showing the effect of mAbs on cholesterol efflux from THP-1 cells.

Figure 6B. Graph showing the effect of mAbs on phospholipid efflux from THP-1 cells.

Figure 7 shows the effect of apoA-I and antibody NDF6F1 on the abundance of ABCAl cells on the surface.

Figure 8 shows the effect of apoA-I and antibodies NDF6F1 and NDF4C2 on the degradation rate of ABCAl.

Figure 9 shows cholesterol efflux from HeLa cells mock-transfected (A) or transfected with ABCAl (B) or ΔPEST-ABCA1 (C).

Figure 10. Shows the effect of antibodies on cholesterol (A) and cholesteryl ester (B) synthesis in macrophages.

RAW 264.7 macrophages were preincubated with the antibodies (final concentration 5 μg/ml) for 18 h at 37° C. Cells were then incubated for 2 h at 37° C. with [3H] acetate and [14X]oleic acid (complexed to BSA). Cells were washed and lipids were extracted and analyzed by TLC. Spots of cholesterol and cholesteryl oleate were identified by standards (Sigma), scraped and counted in a β-counter. *p<0.01 versus no antibody.

Figure 11. Shows the effect of monoclonal antibodies on cholesterol efflux from THP-1 cells.

Cholesterol efflux experiments were conducted as described in the Examples section. Cholesterol efflux is expressed as the percentage of labeled cholesterol moved from cells to medium (i.e. radioactivity in the medium/radioactivity in the medium+radioactivity in the cells). Means±SD of quadruplicate determinations are shown. *p<0.01 versus non-specific IgM.

Figure 12 shows a Western blot using antibodies against ABCAl.

Cells were lysed in RIPA buffer and proteins were separated on a 7.5% SDS-polyacrylamide gel followed by immunoblotting. 1-THP-1 cells; 2-HEK2931 cells transfected with mouse ABCA1; 3-HepG2 cells; 4-CHOP cells.