Serivisi ishinzwe gusesengura poroteyine

Alpha Lifetech irashobora kandi guha abakiriya isesengura rya poroteyine, isesengura rya poroteyine, isesengura rya poroteyine-poroteyine (CO-IP, Western Blot, Chromatin immunoprecipitation assay), hamwe n’imikorere ya poroteyine kugira ngo ihuze ibyifuzo by’abakiriya batandukanye.

TWANDIKIRE

Serivisi ishinzwe gusesengura poroteyine

Alpha Lifetech imaze imyaka myinshi ikora cyane mubushakashatsi bwa poroteyine yubatse urubuga rwiza rwerekana imikorere ya poroteyine, kandi yize uburyo butandukanye bwa tekiniki bwo kugenzura imikoranire ya poroteyine, ishobora kubika ubushakashatsi bwa siyansi cyangwa igihe cy’ubushakashatsi mu gihe itanga ibisubizo bihamye.

Intangiriro Kuri Protein-Protein Imikoranire

Poroteyine nizo molekile zikomeye cyane muburyo ubwo aribwo bwose bwubuzima busubiza amabwiriza yabitswe mubikoresho bya geneti. Kenshi na kenshi, poroteyine zikora akazi kazo zikora nk'abagize uruhare mu guhuza izindi poroteyine, ibyo bikaba bigaragara cyane iyo imikorere ya poroteyine isuzumwe mu buryo nyabwo bw'akagari. Kumenya imikoranire hagati ya poroteyine ningirakamaro cyane mubushakashatsi bwibinyabuzima bwa proteine kugiti cye. Isesengura rya poroteyine yubwoko busanzwe bukurikira:

Co-immunoprecipitation Assay

Ihame ryuburyo bwa immunoprecipitation nuburyo bwo kuvanga poroteyine yagenewe, antibody yihariye, hamwe na poroteyine A / G ya magnetique ya incubation, supernatant ya proteine idafunze ijugunywa hifashishijwe adsorption yamasaro ya magneti kuri rack ya magnetique, kandi amasaro ya magneti yogejwe kugirango akuremo proteine nibindi byanduye keretse bihambiriwe byumwihariko.

Igishushanyo.1 Igishushanyo mbonera cya Co-IP.(Inkomoko yerekana: Coimmunoprecipitation assay - PubMed (nih.gov))

Kuramo hasi

Gukuramo-gukuramo ni ugukosora poroteyine izwi ku isaro rya magneti no kongeramo ibintu bigomba kumenyekana. Eluent yamenyekanye kuri adsorption ya poroteyine zikorana.

Mu 1988, Smith n'abandi. poroteyine ya GST isukuye, yakoreshejwe muburyo bwo gukuramo kandi yitwa Gst gukuramo ni ukuvuga kongeramo tagi ya GST kuri poroteyine yagenewe, gufata poroteyine zikorana binyuze muri GSH, gusobanura guhuza, no kubigenzura binyuze muri WB.

Igishushanyo.2 Igishushanyo mbonera cya GST gukuramo hasi.(Inkomoko yerekana: GST Gukuramo Hasi Kwiga PIF4 Guhuza Vitro - PubMed (nih.gov) )

Gukuramo kwe ni ugukoresha ioni ibyuma nka nikel cyangwa cobalt nkibikoresho, kweza poroteyine binyuze muri chromatografi ya affinity, no gusobanura guhuza binyuze mu bushakashatsi bwa WB kugirango bugenzurwe.

Hiyongereyeho, hariho ADN ikurura (Protein dna binding assay), RNA Gukuramo hasi (Protein rna imikoranire), na molekile nto ikuramo ibice.

ADN Gukuramo (Protein dna binding assay) nuburyo bwa vitro bwo kumenya ADN ihuza poroteyine. Niba poroteyine zihariye zihuza ADN zagaragajwe na WB; Ibice bya ADN bitazwi bihujwe na poroteyine, ibyo bice bya ADN bizwi na MS detection

RNA Gukuramo hasi (Protein rna imikoranire ya assay) nuburyo bwa vitro bwo kumenya RNA ihuza poroteyine. Niba poroteyine zihariye zihuza intego RNA zagaragajwe na WB; Ibice bya RNA bitazwi bihujwe na poroteyine, ibice bya RNA bizwi na MS detection

SM gukuramo (kugabanuka kwa molekile ntoya) irashobora kubona poroteyine zigamije guhuza cyane na molekile ntoya, kandi iyo ihujwe na MS, poroteyine ntoya ya molekile nto irashobora kugenzurwa neza, ishobora kugabanywa muburyo bwa vitro labels hamwe nuburyo bwa biologiya orthogonal.

Ubushakashatsi bwa WB

Ubushakashatsi bwa WB (buzwi kandi nka Western Blot igeragezwa cyangwa Western blot), ibyingenzi nigisubizo cyihariye cya antigen na antibody, ihame ryibanze ni ugutandukanya poroteyine zanduye binyuze muri SDS polyacrylamide gel electrophoreis no kuyimurira mu cyuma gikomeye (nka PVDF membrane, NC membrane) binyuze mu kwimura membrane (kuzunguruka cyangwa igice cyumye). Nyuma yo guhagarika (ukoresheje BSA, buttermilk, nibindi), antibody yibanze ikoreshwa kugirango ihuze proteine, hanyuma antibody ya kabiri ya fluorescein yongeweho nyuma yo koza firime. muguhuza antibody ya kabiri hamwe na antibody yibanze, poroteyine yagenewe irashobora kugaragazwa niterambere ryibara rya substrate hamwe na chemiluminescence. Mubisanzwe bikoreshwa mukumenya niba proteine yihariye igaragarira murugero no gusesengura hafi urwego rwimvugo.

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Kugereranya poroteyine-Poroteyine Uburyo bwo kugereranya

Intungamubiri za poroteyine zisa na Co-IP, zombi zikaba ari zo kumenya ubushakashatsi bwakozwe na poroteyine. Itandukaniro riri hagati yuburyo bubiri nuko Co-IP ivuga ko ibona poroteyine zikorana na poroteyine zizwi, zifite ukuri runaka ariko ntizishobora kwemeza niba poroteyine zikorana zikora mu buryo butaziguye cyangwa butaziguye. Gukuramo hasi ni ugushaka poroteyine itazwi ikorana na poroteyine izwi. Irashobora kwemeza mu buryo butaziguye niba poroteyine izwi ikorana na poroteyine yamenyekanye ariko ntishobora kumenya uko ibintu bimeze muri vivo.

Ikoranabuhanga rya immunoprecipitation rikoreshwa mu gusesengura niba hari imikoranire hagati ya poroteyine ebyiri cyangwa nyinshi, zishingiye ku bushakashatsi bwakozwe na immunoprecipitation, Ugereranije n’ubundi buryo bwo gusesengura imikoreshereze ya poroteyine y’ubuhanga bw’ubushakashatsi (GST Pull-down, Western Blot, Chromatin immunoprecipitation assay, nibindi), ikoranabuhanga rya Co-IP rifite umwihariko wo guhuza no guhuza poroteyine zidasanzwe.

	GST Kumanura	Co-IP	WB
Ibyiza	* Biroroshye gukora * Bikwiranye no kweza poroteyine zitandukanye	* Kuberako muri vivo imikoranire * Irashobora kumenyekana hamwe na poroteyine yo hepfo	* Umwihariko * Ubwiza nubwinshi muri vitro
Imipaka	* Ahari guhuza bidafite akamaro * Ibindi byemezwa byimikoranire birakenewe	* Kwishingikiriza cyane kuri antibodies * Guhuza bidafite akamaro birashobora kubaho	* Bitwara igihe * Biragoye gukoresha kubisesengura binini bya poroteyine
Urugero	* Menya imikoranire * Poroteyine isukuye	* Kwiga transduction Uburyo bw'indwara	Isesengura ryakurikiyeho rya poroteyine-proteyine, proteyine-ADN, hamwe na poroteyine-RNA