Naslov
A preliminary toolkit for rat IgG glycosylation profiling: towards comprehensive rodent animal glyco(proteo)mics

Autori
Habazin, Siniša ; Novokmet, Mislav ; Štambuk, Jerko ; Razdorov, Genadij ; Keser, Toma ; Lauc, Gordan

Vrsta, podvrsta i kategorija rada
Sažeci sa skupova, sažetak, znanstveni

Izvornik
2nd GlycoCom 2018 and 1st Human Glycome Project Meeting book of abstracts / - , 2018, 95-96

Skup
2nd GlycoCom 2018 and 1st Human Glycome Project Meeting

Mjesto i datum
Dubrovnik, Hrvatska, 03.10.2018. - 06.10.2018

Vrsta sudjelovanja
Poster

Vrsta recenzije
Recenziran

Ključne riječi
N-glycosylation ; IgG ; rat ; mass spectrometry ; glycoproteomics

Sažetak
Lab rats have traditionally been favored as experimental animals because, compared to mice now holding primacy as a vertebrate disease model, their physiology and metabolism resembles more closely that of the humans. Since no -omics studies can't be considered complete anymore without introducing glycomic dataset, mouse IgG N-glycosylation is already well explored.The fact that rat serum glycoprofiling has been conducted only recently calls for a thorough examination of normal and pathological rat igG N-glycosylation patterns, encompassing both glycomic and glycoproteomic approach. Here we present an array of protocols for efficient rat IgG isolation using protein L agarose affinity beads, in-gel and in-solution PNGaseF deglycosylation, procainamide labelling of free glycans, purification and PGC enrichment as well as tryptic glycopeptides preparation and RP-SPE cleanup for subclass- specific IgG analysis. Suitable UPLC-FLR and nano-LC-ESI-QqTOF methods were also developed. Pooled serum from specific-pathogen-free Wistar rats was used as a test sample and preliminary results indicate excellent recovery of IgG, but low released glycan and subsequent glycoform diversity pointing out for further method optimization and analysis of larger cohorts originating from different rat stocks and strains. next, we strive to introduce for the first time a high-throughput rat IgG isolation approach using protein L 96-well monolithic plate and eventually compare in detail rat IgG glycosylation with those of mouse and human.

Izvorni jezik
Engleski

Znanstvena područja
Kemija, Biologija, Farmacija, Biotehnologija u biomedicini (prirodno područje, biomedicina i zdravstvo, biotehničko područje)

POVEZANOST RADA