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. 2010 Aug;51(8):2282–2294. doi: 10.1194/jlr.M006759

Fig. 7.

Fig. 7.

TLC immunodetection of lipoprotein-associated individual neutral GSLs from human plasma. GSL-aliquots of extracts from VLDL, LDL, and HDL lipoprotein-fractions and from human (hu) plasma were separated by TLC and analyzed with polyclonal antibodies (pAb) against Lc2Cer, Gb3Cer, Gb4Cer, and nLc4Cer in comparison to Stx1/anti-Stx1-antibody and Stx2/anti-Stx2-antibody. Bound anti-GSL and anti-Stx-antibodies were visualized with alkaline phosphatase conjugated secondary antibodies and BCIP as a substrate. Aliquots applied for anti-Lc2Cer and anti-nLc4Cer immunostains correspond to 12.2 mg and those used for anti-Gb3Cer, anti-Gb4Cer, and the Stx1 and Stx2 overlay assays are equivalent to 18.0 mg of human plasma proteins. GSL aliquots of lipoprotein extracts equal the amounts of 40 µg in the anti-Lc2Cer and anti-nLc4Cer, 60 µg in the anti-Gb3Cer and anti-Gb4Cer, and 120 µg of lipoproteins in the Stx1- and Stx2-overlay assays. The densitometrically determined relative distributions of GSLs between the different lipoproteins were calculated as percentage values and are presented for the antibody-detected GSLs as arabic numerals. The vertical white lines indicate areas of noncontiguous lanes assembled. The GSL structures and backgound information on the employed antibodies and Stxs are provided in Table 1.