Comparison and Optimization of Quantification Methods for Shigella flexneri Serotype 6 O-antigen Containing Galacturonic Acid and Methyl-Pentose

• Published in: International journal of molecular sciences Vol. 22; no. 22; p. 12160
• Main Authors: Raso, Maria Michelina, Vassallo, Oscar, Micoli, Francesca, Giannelli, Carlo
• Format: Journal Article
• Language: English
• Published: Switzerland MDPI AG 10.11.2021; MDPI
• Subjects: Antigens; Antimicrobial resistance; Calibration; Colorimetry; D-Galactosamine; Design of experiments; Diarrhea; Dische colorimetric method; Glucose; Hexuronic Acids - chemistry; Hexuronic Acids - isolation & purification; HPAEC-PAD; Immune response; Methods; Morbidity; O Antigens - chemistry; O Antigens - isolation & purification; Pentose; Pentoses - chemistry; Pentoses - isolation & purification; polysaccharide; Polysaccharides; Public health; Quality management; quantification; Rhamnose; S. flexneri O-antigen; Shigella; Shigella flexneri - chemistry; Statistical analysis; vaccine; Vaccine development; Vaccines; methyl-pentose; vaccine; quantification; polysaccharide; Dische colorimetric method; design of experiment; S. flexneri O-antigen; HPAEC-PAD; galacturonic acid
• ISSN: 1422-0067; 1661-6596; 1422-0067
• DOI: 10.3390/ijms222212160

is a leading diarrheal cause of morbidity and mortality worldwide, especially in low- and middle-income countries and in children under five years of age. Increasing levels of antimicrobial resistance make vaccine development an even higher global health priority. serotype 6 is one of the targets of many multicomponent vaccines in development to ensure broad protection against . The O-antigen (OAg) is a key active ingredient and its content is a critical quality attribute for vaccine release in order to monitor their stability and to ensure appropriate immune response. Here, the optimization of two methods to quantify 6 OAg is reported together with the characterization of their performances. The optimized Dische colorimetric method allows a tenfold increment of the sensitivity with respect to the original method and is useful for fast analysis detecting selectively methyl-pentoses, as rhamnose in 6 OAg. Also, a more specific HPAEC-PAD method was developed, detecting the dimer galacturonic acid-galactosamine (GalA-GalN) coming from 6 OAg acid hydrolysis. These methods will facilitate characterization of 6 OAg based vaccines. The colorimetric method can be used for quantification of other polysaccharide containing methyl-pentoses, and the HPAEC-PAD could be extended to other polysaccharides containing uronic acids.