Synthesis of reference standards to enable single cell metabolomic studies of tetramethylrhodamine-labeled ganglioside GM1.
Ganglioside GM1 and its seven potential catabolic products: asialo-GM1, GM2, asialo-GM2, GM3, Lac-Cer, Glc-Cer and Cer, were labeled with tetramethylrhodamine (TMR) to permit ultra-sensitive analysis using laser-induced fluorescence (LIF) detection. The preparation involved acylation of the homogenous C(18)lyso-forms of GM1, Lac-Cer, Glc-Cer and Cer with the N-hydroxysuccinimide ester of a beta-alanine-tethered 6-TMR derivative, followed by conversion of these labeled products using galactosidase, sialidase, and sialyltransferase enzymes. The TMR-glycolipid analogs produced are detectable on TLC down to the 1 ng level by the naked eye. All eight compounds could be separated within 4 min in capillary electrophoresis where they could be detected at the zeptomole (ca. 1000 molecule) level using LIF.
Larsson
E Andreas
EA
Carlsberg Laboratory, Valby-Copenhagen DK-2500, Denmark.
Olsson
Ulf
U
Whitmore
Colin D
CD
Martins
Rita
R
Tettamanti
Guido
G
Schnaar
Ronald L
RL
Dovichi
Norman J
NJ
Palcic
Monica M
MM
Hindsgaul
Ole
O
D000818:Animals; D002417:Cattle; D019075:Electrophoresis, Capillary; D005456:Fluorescent Dyes / Q000737:chemistry*; D005677:G(M1) Ganglioside / Q000031:analogs & derivatives* / Q000302:isolation & purification / Q000378:metabolism*; D005678:G(M2) Ganglioside / Q000737:chemistry / Q000378:metabolism; D005679:G(M3) Ganglioside / Q000302:isolation & purification; D005732:Gangliosides / Q000302:isolation & purification; D009439:Neuraminidase / Q000378:metabolism; D012015:Reference Standards; D012235:Rhodamines / Q000737:chemistry*
D005456:Fluorescent Dyes; D005679:G(M3) Ganglioside; D005732:Gangliosides; D012235:Rhodamines; D005678:G(M2) Ganglioside; C038244:ganglio-N-triaosylceramide; D005677:G(M1) Ganglioside; C005358:tetramethylrhodamine; C018835:asialo GM1 ganglioside; D009439:Neuraminidase