Ni izihe nyungu zo gukoresha ibikoresho bya BCA?

IRIBURIRO RWA BCA ProtenIn Assay

BCA (bicinchonic acide) poroteyine ifata tekinike yingirakamaro mubyerekezo byubushakashatsi bwibinyabuzima na kuvura selire. Yorohereza umubare wa poroteyine yibanda ku cyitegererezo. Ubu buryo bukora ku ihame rya poroteyine - chelation yumuringa, hakurikiraho ibara rya tapimetric yagabanijwe intungamubiri. Icyamamare cyacyo gituruka ku bikorwa byayo byagutse hirya no hino kuri poroteyine nyinshi, harimo n'abafite inkingi nka surpacts, bituma habaho guhitamo abakora n'abatanga mu murima.

Guhuza hamwe no guhagarika

Gukemura Ingero Zigoye

Imwe mu nyungu zigaragara za poroteine ​​ya BCA ni uguhuza na Cargengano. Bitandukanye nubundi buryo bwinshi bwo kugereranya poroteyine, BCA ifatanije irashobora kwakira kugeza 5% surfactacts zigenda neza murwego. Iyi mikorere ni ingirakamaro cyane kubashakashatsi nabakora bakorana ningero zigoye, aho byakoreshwaga kenshi mugukemura proteyine. Kubikorwa byo kuvura selire, uku guhuza byemeza ko umubare wa poroteyine ukomeza kuba ubwukuri nubwo hariho ibi bintu bibangamiye.

Imikorere mubihe bitoroshye

BCA ifata ibisubizo byizewe nubwo bigoye. Imyitwarire yacyo irwanya itandukaniro muri poroteyine, nka aside aside acino itandukanya itandukaniro ninyungu zuruhande, zigabanya proteyine - Kuri - Guhinduranya Guhinduka. Ubuswa nibyingenzi kubatanga ibicuruzwa bakeneye ibisubizo bihoraho kandi byororoka kugirango bikomeze - Ibipimo byiza muri poroteyine isaba.

Kugabanya igihe cyo gushiraho

Imikorere hamwe no kwikuramo - Ibipimo byubusa

Intangiriro yo Kwirukana - Ibipimo bya poroteyine kubuntu muri bca assay ibikoresho bigabanya cyane umwanya wo gusuzuma kugeza kuri 80%. Aya mahame yagenwe yagenewe kwishyira hamwe kwagaciro hamwe na pipettes nyinshi, kwemerera abashakashatsi nabakora kugirango bihutishe gahunda yo gusuzuma batabangamiye ukuri. Kubikorwa byo kuvura akagari, aho - Ubushakashatsi bwumva burasanzwe, ubu buryo bwo gukora ntagereranywa.

Umukozi woroshye

Iyi gahunda yo kwerekana kugabanya icyifuzo cyo gusubiranamo intoki, kitari igihe - Gukoresha ariko nanone bikunda amakosa. Mugukuraho izi ntambwe, muri BCA Assora Guhinduka, kwemerera abatanga isoko guhura na gahunda yo gutanga no kuzamura umusaruro muburyo bwa laboratoire.

UMURYANGO W'INGENZI

Gupima neza muburyo bwa poroteyine

Umuyoboro wa BCA Engeat Erekana igisubizo kimwe kuri poroteyine zitandukanye ugereranije nizuba - guhuza ibisasu nkimyitozo ya Bradford. Ubuhungiro bugerwaho no kwitondera ugereranije numubare wa Peptade Kubakora ibigize uruhare mugutanga abaganga b'udushinga, ibi bituma hafatwa neza ibipimo nyabyo ahantu heza hatandukanye, kuzamura icyizere mu bikorwa byo gusuzuma ibisubizo.

Kunoza Guhuza Guhoraho

Mugutanga ibisubizo bihamye utitaye ku gutandukana gutandukanya poroine, bca ingirakamaro ni ntagereranywa kubatanga isoko basaba umubare nyawo kugirango ugenzure neza nibipimo ngenderwaho. Uku guhuzagurika guhindura amakuru menshi yizewe, ni ngombwa mugutera gukora ubushakashatsi mubuvuzi bwa selire nibikorwa bifitanye isano.

Urwego rwo kumenya no kumenya

Kwiyumvisha cyane kwibanda

BCA ifata urwego rwo hejuru rwintangarugero, ishoboye gutahura proteine ​​kwibandaho nka 0.5 μg / ml, no kubungabunga umurongo kugeza kuri 1.5. Iyi mibare yagutse ituma bikwiranye nibisabwa bitandukanye, ukurikije ubushakashatsi bwibanze ku iterambere ryateye imbere. Ibitekerezo nkibi ni ingenzi mugihe uhanganye numunota wa poroteyine mubuvuzi bwumuvugizi nubushakashatsi bwa farumasi.

Ibipimo byiza byo gutahura

Gutahura amabara kuri 562 NM iremeza igihombo kinini cyibimenyetso (munsi ya 10%), kwemerera abakora nabatanga ibicuruzwa kugirango bagere ku mibare myiza. Uku kuri kwukuri ni ngombwa mugutezimbere ibicuruzwa byiza bya selire, aho kwibanda kwa poroteyine bishobora kugira ingaruka muburyo butaziguye.

Koroshya gukoresha muburyo bwo gusuzuma inzira

UBURYO

BCA Issay Ibiranga inzira zoroheje, zingirakamaro kubakoresha bashya hamwe nabanyamwuga bamenyereye. Ibyiteguye - Kuri - Koresha reagen hamwe na Porotokole itaziguye igabanya amakosa yabaga, bigatuma habaho abumva mugari, harimo abayikora hamwe nuburyo bushya hamwe nuburyo bwa poroteyine.

Umukoresha - Igishushanyo cya Gicuti

Yaremewe uburyo bworoshye bwo gukoresha mubitekerezo, muri BCA igabanya inzitizi za tekiniki, koroshya amahugurwa byoroshye kandi urwango byihuse mububiko bwa laboratoire. Uku kuboneka bifasha ibikorwa byabatanga ibicuruzwa bakeneye gukomeza kwinjiza byinshi utabangamiye ukuri.

Ukuri no guhuzagurika

Ibisubizo byizewe mubushakashatsi

Umufuka wa BCA utanga ibisubizo byizewe, birangwa na poroteyine nke - kuri - gutandukana kwa proteine. Ubu buryo buhoraho ni ingenzi ku bushakashatsi busaba ibipimo byasubiwemo igihe, nk'ibyo ku buvuzi bwa selire, aho umubare munini wa poroteyine ari umwanya munini wo kubyara no kwemeza ibisubizo.

Imibare na kalibrasi

Imibereho iringanizwa BSA n'ibigo by'igihugu cy'ibipimo ngenderwaho (Nist) biremeza ko BCA ifata umurongo usanzwe kandi woroha. Iyi kalibrasi ningirakamaro kubakorera hamwe nabatanga isoko bigamije gutanga hejuru - ubuziranenge, ibicuruzwa bisanzwe.

Ibice bya tekinike byuburyo bwa BCA

Uburyo bwa Bioniyeli

Uburyo bwa BCA bushingiye ku kugabanya CU²⁺Kuri Cu⁺na poroteyine muburyo bwa alkaline, ikurikiwe no gushiraho uruganda rwumuhengeri hamwe na BCA, ikibera nkigipimo cyamabara. Ibi byombi - INTAMBWE Itanga ishingiro ryizewe rya Proteines, rikoreshwa mubushakashatsi butandukanye, harimo iterambere ryibicuruzwa bya selire.

Ibyiza birenze ibindi bishakiye

Ugereranije nuburyo nka Bradford isaba, uburyo bwa BCA itanga uburyo bwinshi bitewe no guhuza kwagutse no guhuza ibintu bitandukanye hamwe nuburyo buhoraho muburyo butandukanye bwa poroteyine. Ibi bituma habaho guhitamo ababikora nabatanga ibicuruzwa bisaba ibisubizo bivuguruzanya kandi byizewe.

Kugereranya na Bradford Assum

Itandukaniro ryingenzi muburyo bukoreshwa

Mugihe BCA NA BRADFORD isaba cyane ikoreshwa cyane kuri proteine, ziratandukanye cyane muburyo bwabo. Bradford ifatanije kuri doye - proteine ​​irahuza, ishobora kuba idahuye muburyo butandukanye bwa poroteyine. Ibinyuranye n'ibyo, Peptay's Peptay Bond - Gutahura bishingiye ku gitsina gatanga uburinganire kandi bwizewe, cyane cyane kubakora n'abaguzi bibanze ku buvuzi bwa selire n'imivugo.

Ibyiza bya BCAS Assay

Guhuza kwa BCA bihuye nibikoresho byagutse, imbaraga zagutse, hamwe nakazi koroshe biha inkombe kubisabwa bisaba ubu bushobozi bwihariye. Guhinduranya kwayo bituma bikwiranye nimikorere yagutse, ikintu gikomeye kubatanga ibicuruzwa bitanga ibicuruzwa kubikenewe byubushakashatsi bitandukanye.

Umwanzuro n'ibyifuzo

Impamvu BCA ikunzwe

Igitero cya Koroupy gifatanije no guhuza n'ibikoresho byabitswe, byagabanijwe igihe cya SETUP, kimwe cya proterine cyongerewe ubumwe, kandi ukuri. Iyi mico ituma bikwiranye cyane kubisabwa mubuvuzi bwa selire, gutanga ibisubizo byizewe kandi bihamye. Kubakora n'ababitanga, BCA Assay atanga uburyo bwiringirwa bwo kubuza - Umubare ntarengwa wa poroteyine, ni ngombwa mugukomeza ibipimo ngenderwaho no gushyigikira iterambere ryibicuruzwa.

Bluekit tanga ibisubizo

Ubururu bwubururu mugutanga ibisubizo byambere kuri poroteine ​​ya bca poroteine, bidoze kuzuza ibyifuzo byumushakashatsi, abakora, nabatanga isoko. Ibikoresho byacu byateguwe kubikorwa byoroshye, bigatanga poroteyine yizewe kandi ihamye ya poroteyine hejuru ya porogaramu nini. Hamwe na Bluekit, urashobora kongera imikorere yawe no kuba ubwukuri, neza ko wujuje ubuziranenge bwo hejuru mubushakashatsi bwo kuvura akagari no guteza imbere ibicuruzwa. Reka Bluekit ube umufatanyabikorwa wawe wizewe kuri poroteyine yawe yose asaba ibikenewe.