Study on the quality control of cell therapy products. Determination of N-glycolylneuraminic acid incorporated into human cells by nano-flow liquid chromatography/Fourier transformation ion cyclotron mass spectrometry.
N-Glycolylneuraminic acid (NeuGc), an acidic nine-carbon sugar, is produced in several animals, such as cattle and mice. Since human cells cannot synthesize NeuGc, it is considered to be immunogenic in humans. Recently, NeuGc contamination was reported in human embryonic stem cells cultured with xenogeneic serum and cells, suggesting that possibly NeuGc may harm the efficacy and safety of cell therapy products. Sialic acids have been determined by derivatization with 1,2-diamino-4,5-methylenedioxybenzene (DMB) followed by liquid chromatography/mass spectrometry (LC/MS) and liquid chromatography/tandem mass spectrometry (LC/MS/MS); however, the limited availability of cell therapy products requires more sensitive and specific methods for the quality test. Here we studied the use of nano-flow liquid chromatography/Fourier transformation ion cyclotron resonance mass spectrometry (nanoLC/FTMS) and nanoLC/MS/MS for NeuGc-specific determination at a low femtomole level. Using our method, we found NeuGc contamination of the human cell line (HL-60RG cells) cultured with human serum. Our method needs only 2.5x10(3) cells for one injection and would be applicable to the determination of NeuGc in cell therapy products.
Hashii
Noritaka
N
Division of Biological Chemistry and Biologicals, National Institute of Health Sciences, 1-18-1 Kamiyouga, Setagaya-ku, Tokyo 158-8501, Japan.
Kawasaki
Nana
N
Nakajima
Yukari
Y
Toyoda
Masashi
M
Katagiri
Yoko
Y
Itoh
Satsuki
S
Harazono
Akira
A
Umezawa
Akihiro
A
Yamaguchi
Teruhide
T
D002138:Calibration; D002462:Cell Membrane / Q000737:chemistry; D064987:Cell- and Tissue-Based Therapy / Q000592:standards*; D002853:Chromatography, Liquid / Q000379:methods*; D017357:Cyclotrons; D005583:Fourier Analysis; D018922:HL-60 Cells; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods*; D019158:N-Acetylneuraminic Acid / Q000032:analysis / Q000737:chemistry; D036103:Nanotechnology / Q000379:methods*; D009438:Neuraminic Acids / Q000032:analysis* / Q000737:chemistry; D011786:Quality Control; D013347:Subcellular Fractions / Q000737:chemistry
D009438:Neuraminic Acids; C032592:N-glycolylneuraminic acid; D019158:N-Acetylneuraminic Acid