A specific sample preparation method was developed for this work, using 2,5-dihydroxybenzoic acid
 Nonstandard abbreviations: LSD, lysosomal storage disease; FOS, free oligosaccharides; MS, mass spectrometry; AGU, aspartylglucosaminuria; ANTS, 2-aminonaphthalene trisulfone; GlcNAc, N-acetylglucosamine; MALDI-TOF/TOF, MALDI-time-of-flight/time-of-flight; DHB, 2,5-dihydroxybenzoic acid
; TFA, trifluoroacetic acid; Asn, asparagine; ML, mucolipidosis; GNPT, GlcNAc phosphotransferase.
 Nonstandard abbreviations: CDG, congenital disorder of glycosylation; TIEF, isoelectric focusing of plasma transferrin; HUS, hemolytic uremic syndrome; MS, mass spectrometry; MSn, multistage MS; GDP-Fuc, GDP-fucose; PNGase F, peptide-N4-(acetyl-[beta]-glucosaminyl)-asparagine amidase; DHB, 2,5-dihydroxybenzoic acid
; DMA, N,N-dimethylaniline; NeuNAc, complex N-glycans with at least 1 antenna without a terminal sialic acid; GlcNAc, N-acetyl glucosamine; Gal, galactose; COG, conserved oligomeric Golgi.
About 2-mg samples were dissolved in 1 mL of hexafluoroisopropanol (HFIP), and 2,5-dihydroxybenzoic acid
(DHB), indoleacrylic acid, or 2-(4-hydroxyphenylazo)-benzoic acid (HABA) (0.5 M in HFIP as solvent) were used as matrix.
We purchased the matrix, 2,5-dihydroxybenzoic acid
(DHB), trifluoroethanol, sodium hydroxide, CHAPS, Tris-HCl, sodium pyrophosphate, EDTA, and EGTA from Aldrich; chloroform, iodomethane, and sodium chloride from EM Science; dithiothreitol (DTT) and iodoacetamide (IAA) from Bio-Rad Laboratories; ammonium bicarbonate from Mallinckrodt Chemical Company; and acetonitrile from Fisher Scientific.
 Nonstandard abbreviations: CDC, congenital disorders in glycosylation; MS, mass spectrometry; ALP, alkaline phosphatase; AFP, [alpha]-fetoprotein; DHB, 2,5-dihydroxybenzoic acid
; LR, likelihood ratio; AUC, area under the curve; G1cNAc, N-acetyl-glucosamine; APTT, activated partial thromboplastin time.
Finally, each sample was mixed with 5 [micro]L of 10 g/L 2,5-dihydroxybenzoic acid
(DHB) as a matrix.
SA was selected as the MALDI-TOF MS matrix of choice for this investigation because other standard matrices used in protein analysis, [alpha]-cyano-4-hydoxycinnamic acid and sDHB [a 9:1 (by weight) mixture of 2,5-dihydroxybenzoic acid
and 5-methoxysalicylic acid], did not yield intense spectra for hCG[beta]cf (data not shown).
An alternative matrix that has worked well is 10 g/L 2,5-dihydroxybenzoic acid
in 300-500 mL/L acetonitrile containing 1 mL/L trifluoroacetic acid.