Kayayyaki

Sabon babban aikin ruwa chromatography kima don glycosyltransferases dangane da rarrabuwa tare da acid anthranilic da gano haske.

An haɓaka ƙididdiga ta amfani da sinadarai na musamman na 2-aminobenzoic acid (2AA; anthranilic acid, AA) don auna ayyukan biyu na β1-4 galactosyltransferase (GalT-1) da α2-6 sialyltransferase (ST-6) ta ruwa mai girma. chromatography (HPLC) tare da gano haske mai haske (Anumula KR. 2006. Ci gaba a cikin hanyoyin samar da hasken wuta don babban aiki na chromatographic ruwa na glycoprotein carbohydrates. Anal Biochem. 350: 1-23).An yi amfani da N-Acetylglucosamine (GlcNAc) da N-acetyllactosamine a matsayin masu karɓa da uridine diphosphate (UDP) -galactose da cytidine monophosphate (CMP) -N-acetylneuraminic acid (NANA) a matsayin masu ba da gudummawa ga GalT-1 da ST-6, bi da bi.An yi wa samfuran Enzymatic alama a cikin wurin tare da AA kuma an raba su daga abubuwan da ke kan ginshiƙi na TSKgel Amide 80 ta amfani da yanayin-lokaci na yau da kullun.An ƙayyade raka'o'in Enzyme daga wurare mafi girma ta hanyar kwatantawa tare da ƙa'idodin da aka haɗa tare da Gal-β1-4GlcNAc da NANA-α2-6 Gal-β1-4GlcNAc.Linearity (lokaci da ƙaddamarwar enzyme), daidaito (intra- da interassay) da sake sakewa don ƙididdigar an kafa su.An gano ƙididdigar suna da amfani wajen saka idanu akan ayyukan enzyme yayin keɓewa da tsarkakewa.Gwaje-gwajen sun kasance masu hankali sosai kuma an yi daidai da ko mafi kyau fiye da ma'aunin tushen sukari na rediyoaktif na gargajiya.Hakanan za'a iya amfani da tsarin tantancewa don auna ayyukan wasu masu canja wuri, muddin masu karɓar carbohydrate sun ƙunshi ƙarshen ragi don lakabi.An haɓaka ƙima don masu karɓar glycoprotein ta amfani da IgG.An ɓullo da ɗan gajeren hanyar bayanin martaba na HPLC don rabuwa da IgG glycans (biantennary G0, G1, G2, mono- da disiaylated), wanda ya sauƙaƙe ƙaddamar da ayyukan GalT-1 da ST-6 cikin sauri.Bugu da ƙari, wannan hanyar bayanin martaba yakamata ya zama mai amfani don sa ido kan canje-canje a cikin IgG glycans a cikin saitunan asibiti.