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. 2020 May 21;21(10):3632. doi: 10.3390/ijms21103632

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© 2020 by the author.

Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).

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Characterization of monoclonal antibodies generated from a Gb4Cer-VLCFA-immunized mouse. (A) Reactivity of monoclonal antibodies (mAbs) PA4.2, PA5, and PA7 against several GSLs, evaluated by ELISA. The wells of a 96-well microtiter plate were coated with 500 ng of each GSL and then reacted with each mAb at a concentration of 2 μg/mL. The structures of the GSLs are summarized in Table 3. (B) Reactivity of mAb PA7 against various concentrations of Gb4Cer (Gb4) and Gb3Cer (Gb3), evaluated by ELISA. (C) Thin-layer chromatography (TLC)-immunostaining of Gb4Cer using PA7. The double band found in Gb3Cer is due to the difference in the lipid structure. Left panel, orcinol-H₂SO₄ staining. Right panel, immunostaining with PA7. St, standard glycosphingolipids mixture; Gb4, Gb4Cer; Gb3, Gb3Cer; CMH, ceramide monohexoside; CDH, ceramide dihexoside.