GlycoDigest – Exoglycosidase Brief Guide

IgG Fc Glycopeptides by LC-MS Immunoglobulin G antibodies

MDCK HCP ELISA Kit

Resume

Glycosphingolipids (GSLs) from two Madin-Darby canine kidneys (MDCK) cell sublines, an epithelial cell line, were characterized by t.l.c., antibody overlay, and mass spectrometry. The main characteristic that distinguishes the two strains of MDCK cells is their transepithelial electrical resistance which is typically on the order of 3000 ohms.cm2 for the strain I cells and 100 ohms.cm2 for strain II cells. Cells from strains I and II were equally rich in glycolipids, the cell GSL/phospholipid ratio being 0.04.

However, although the phospholipid patterns were identical, the GSLs showed striking differences, with each cell strain expressing appreciable amounts of GSLs that were not found in the other strain. Both cell types possessed neutral GSLs with one, two, or three carbohydrate moieties. Monoglycosylceramide represented 50% of the total GSL in each strain. However, while in strain I cell more than 90% of this monoglucosylceramide was monoglucosylceramide, in strain II cells more than 90% consisted of monogalactosylceramide.

In addition, MDCK strain II cells selectively expressed GSLs belonging to the globoid series (26% of their neutral GSLs), including globoside and Forssman’s antigen, a globoside derivative. The strain I MDCK cells, on the other hand, expressed another series of GSLs with 4-7 carbohydrate residues characterized by the common sequence Hex-HexNAc-Hex-Hex-Cer. The presence of two fucosylated GSLs in these series was established. Both strain I and II MDCK HCP ELISA Kit contained negatively charged GSL, the main component of which was the ganglioside GM3. Strain II MDCK cells also expressed sulfatide, the sulfated derivative of galactosylceramide.

Origin of the cell line

Canine cocker spaniel kidney

Cell line description

From the kidney of a normal adult female Cocker Spaniel in 1958 by SH Madin and NB Darby (Madin Darby Canine Kidney). Supports the growth of a wide range of animal viruses: VSV (Indiana strain), Infectious Canine Hepatitis, Vaccinia, Coxsackie B5, Adeno and reoviruses, SVEV.

Application

Virus studies: SVEV, VSV, Vaccinia, adeno and reoviruses, Coxsackie B5 et al

Culture medium

EMEM (EBSS) + Glutamine 2mM + 1% Non-Essential Amino Acids (NEAA) + 10% Fetal Bovine Serum (FBS).

Subculture routine

Split subconfluent cultures (70-80%) 1:3 to 1:10, ie, seed at 1-3×10,000 cells/cm2 using 0.25% trypsin/EDTA; 5% CO2; 37°C Cells adhere strongly and require at least 2 washes with PBS before adding trypsin/EDTA.

Other notes

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