Journal: Journal of Lipid Research

Article Title: ACAT2 contributes cholesteryl esters to newly secreted VLDL, whereas LCAT adds cholesteryl ester to LDL in mice

doi: 10.1194/jlr.m500018-jlr200

Figure Lengend Snippet: Fig. 1. Representative quantitative apolipoprotein B (apoB) West- ern blot of liver perfusate from an ACAT2-deficient/LDL receptor- deficient (ACAT2/ LDLr/) mouse fed a 0.02% cholesterol diet. Standards and time points were prepared and immunoblotted us- ing a polyclonal antibody raised against mouse apoB-100 as de- scribed in Materials and Methods. Lanes 1–6 represent serial dilu- tions of standards, with amounts of either apoB-100 or apoB-48 indicated at bottom. Lanes 7–13 represent time points from 3 h liver perfusion, with the time points indicated at bottom.

Article Snippet: Electrophoresis was carried out for 6 h at 40 V, and by guest, on A pril 14, 2015 w w w .jlr.org D ow nloaded from Lee et al. VLDL and LDL cholesteryl esters from ACAT2 and LCAT 1207 the gel was then blotted onto nitrocellulose for 2 h at 100 V. The blot was blocked in 3% nonfat dry milk in TBST (150 mM NaCl, 20 mM Tris, pH 7.4, and 0.05% Tween 20) for 1 h and then incubated for 2 h at room temperature in a rabbit anti-mouse apoB polyclonal antibody (Bio-Rad) diluted 1:5,000 in primary antibody buffer (TBST with 0.5 mg/ml MgCl 2 ).

Techniques:

Journal: Journal of Lipid Research

Article Title: ACAT2 contributes cholesteryl esters to newly secreted VLDL, whereas LCAT adds cholesteryl ester to LDL in mice

doi: 10.1194/jlr.m500018-jlr200

Figure Lengend Snippet: Fig. 2. ApoB accumulation rates in liver perfusate of ACAT2/ LDLr/ (aarr) and LDLr/ (rr) mice fed either a high-cholesterol (HC) or a low-cholesterol (LC) diet. ApoB-100 and apoB-48 bands from immunoblots were quantified as described in Materials and Methods, and accumulation rates were calculated. Values represent mean total apoB (apoB-100 plus apoB-48) accumulation rates SEM. n 4 for all groups except the low-cholesterol LDLr/ mice, which had n 3.

Article Snippet: Electrophoresis was carried out for 6 h at 40 V, and by guest, on A pril 14, 2015 w w w .jlr.org D ow nloaded from Lee et al. VLDL and LDL cholesteryl esters from ACAT2 and LCAT 1207 the gel was then blotted onto nitrocellulose for 2 h at 100 V. The blot was blocked in 3% nonfat dry milk in TBST (150 mM NaCl, 20 mM Tris, pH 7.4, and 0.05% Tween 20) for 1 h and then incubated for 2 h at room temperature in a rabbit anti-mouse apoB polyclonal antibody (Bio-Rad) diluted 1:5,000 in primary antibody buffer (TBST with 0.5 mg/ml MgCl 2 ).

Techniques: Western Blot

Journal: Journal of Lipid Research

Article Title: ACAT2 contributes cholesteryl esters to newly secreted VLDL, whereas LCAT adds cholesteryl ester to LDL in mice

doi: 10.1194/jlr.m500018-jlr200

Figure Lengend Snippet: Fig. 4. Negative stain electron microscopy of apoB-containing lipo- proteins isolated from liver perfusion and plasma of knockout mice fed a 0.02% cholesterol diet. Negative stain electron microscopy was carried out as described in Materials and Methods. All images are shown at a magnification of 90,000 . A: VLDL isolated from ACAT2/ LDLr/ mouse liver perfusion. B: VLDL isolated from ACAT2/ LCAT/ LDLr/ mouse liver perfusion. C: VLDL iso- lated from ACAT2/ LDLr/ mouse plasma. D: VLDL isolated from ACAT2/ LCAT/ LDLr/ mouse plasma. E: LDL isolated from ACAT2/ LDLr/ mouse plasma. F: LDL isolated from ACAT2/ LCAT/ LDLr/ mouse plasma.

Article Snippet: Electrophoresis was carried out for 6 h at 40 V, and by guest, on A pril 14, 2015 w w w .jlr.org D ow nloaded from Lee et al. VLDL and LDL cholesteryl esters from ACAT2 and LCAT 1207 the gel was then blotted onto nitrocellulose for 2 h at 100 V. The blot was blocked in 3% nonfat dry milk in TBST (150 mM NaCl, 20 mM Tris, pH 7.4, and 0.05% Tween 20) for 1 h and then incubated for 2 h at room temperature in a rabbit anti-mouse apoB polyclonal antibody (Bio-Rad) diluted 1:5,000 in primary antibody buffer (TBST with 0.5 mg/ml MgCl 2 ).

Techniques: Staining, Electron Microscopy, Isolation, Clinical Proteomics, Knock-Out