Miedzo yembiriso yemethylene blue: kuverenga masero mapenyu zvichienzaniswa nekuyera nguva yekufema

Machubhu maviri ari patafura yerabhoritari. Ose ari maviri ane ruvara rwebhuruu. Mune imwe, masero ebhuruu anoonekwa nemaikorosikopu anoreva mbiriso yakafa. Mune imwe, chubhu inoshanduka kubva pabhuruu ichishaya ruvara inoreva kuti mbiriso ipenyu uye iri kufema. Dhayi rimwe chete, asi zvabuda zvinoreva zvinhu zvakapesana. Ukavhiringidza miedzo iyi miviri, mhedziso dzose dzinopinduka.

Musiyano uyu ndiwo musimboti wechinyorwa chose. Methylene blue idhayi reredox: kana raitwa okisidhesheni rine ruvara rwebhuruu, kana raderedzwa rinova leucomethylene blue isina ruvara. Kuverenga masero mapenyu kunoshandisa kemisitiri iyi mukuongorora kupfupi nemaikorosikopu. Muedzo wekufema unoishandisa mukuita kwemusanganiswa wose kunoyerwa nguva. Mumwe nomumwe unoda maitiro awo, miedzo yawo yekuenzanisa, uye kunyatsongwarira pakududzira zvabuda.

Kuverenga masero mapenyu: penda kwenguva pfupi, woongorora nekukurumidza

Maitiro anoshandiswa mukubika doro ndiwo akanyanya kuiswa pamwero mumwe. Gadzira 0.01% w/v methylene blue mu2% sodium citrate, woisanganisa 1:1 nemusanganiswa wembiriso iri mumvura, uye ongorora musanganiswa mushure menguva pfupi yakatarwa. Nguva dzakabudiswa dzinosiyana kubva pa1 kusvika pa2 maminitsi mubhuku reBrewing Science Institute kusvika pamaminitsi anenge 5 mumaitiro etsvakurudzo, ukuwo nzira yeOIV yekuongorora tupenyu tudiki muwaini ichida kuti kuongorora kuitwe mukati memaminitsi 10. Dhayi mumusanganiswa wekuverengera rinozopedzisira rava pedyo ne0.005%. Isa musanganiswa mukamuri yekuverengera yeNeubauer yakavandudzwa, uye pakukudza kunenge 400x isa masero ane bhuruu yakasviba muboka reasina upenyu uye asina kupendwa muboka remapenyu. Masero ane bhuruu yakareruka anoverengwa semapenyu munzira yeOIV, zvinototaridza dambudziko rekutonga kunosiyana nemuongorori rinotsanangurwa pasi apa.

Huwandu hwemasero pavhoriyamu hunobva pachimiro nezviyero zvekamuri. Kana sikweya yose yepakati ye1 mm yaverengwa, masero pa mL akaenzana neN inopetwa nechiyero chekudiluta uye ne10^4, apo N iri huwandu hwakaverengwa munzvimbo iyoyo, sezvinotsanangurwa munzira dzeASBC dzembiriso. Chikamu chemasero mapenyu chiri nyore kuwana: masero asina kupendwa anopatsanurwa nemasero ose, zvopetwa ne100. Verenga masero anenge 400 kuti ufungidzire chikamu chemapenyu, uchitevedzera tsvakurudzo yeFEMS yekuenzanisa nzira mukubika doro nemaitiro eOIV. Shandisa mutemo mumwe wemiganhu nguva dzose, somuenzaniso kusanganisira masero ari pamitsetse yepasi neyekuruboshwe yegridhi uye kusaverenga ari pamitsetse yepamusoro neyekurudyi.

Mabukira anoda mutemo kuverenga kusati kwatanga. Mutemo unowanzoshandiswa unoverenga bukira sesero rakazvimirira kana rakura kusvika pahafu yehukuru hwesero raamai, uye unoriverenga pamwe chete naamai kana riri diki pane izvozvo. Chero mutemo wasarudzwa, ushandise pasikweya imwe neimwe uye uunyore papepa rekuverengera kuitira kuti munhu anotevera audzokorore.

Pepa rekuverengera (dhinda iri)

Pepa iri rakagadzirwa zvakananga nokuda kwechinyorwa chino. Rikopere mubhuku rekunyora kana kudhinda rimwe pasampuli imwe neimwe.

Chikamu chemasero mapenyu (%) = 100 x asina kupendwa / (asina kupendwa + ane bhuruu yakasviba). Masero ose pa mL = N x chiyero chekudiluta x 10^4, N ichibva munzvimbo yakatarwa yekamuri. Nyora nguva yekupenda pedyo nezvabuda, nokuti nhamba yechikamu chemasero mapenyu isina nguva yayo haigoni kudzokororwa zvakavimbika.

Nei nhamba dzechikamu chemasero mapenyu dzichisiyana

Nguva ndiyo yokutanga inokanganisa, uye inogona kukanganisa munzira mbiri. Masero ari kuita mabasa emetabolism anoderedza dhayi, saka kuongorora nekukasika kunogona kubata masero mapenyu asati apedza kurideredza woaisa muboka reakafa zvisizvo. Asi kugara mudhayi kwenguva refu pachako kune chepfu uye kunokonzera masero ebhuruu akakuvadzwa zvechokwadi, zvichipa kupendwa kunoratidza kufa nenzira inotsausa. Naizvozvo maitiro ekudzidzisa murabhoritari anoti kuverenga kuitwe mukati memaminitsi 5, sezvakanyorwa mumaitiro ekuWisconsin ekudzidzisa nezvembiriso. Tsananguro yakangwarira ndeyokuti kuverenga nemethylene blue kunoenderana nenguva, saka sarudza nguva imwe pfupi uye uichengetedze yakafanana pasampuli imwe neimwe iri muboka.

Sampuli dzine chikamu chidiki chemasero mapenyu dzine kumwe kurerekera kwechipiri. Nzira yakaiswa pamwero mumwe inoyambira kuti methylene blue inogona kupa fungidziro yechikamu chemasero mapenyu yakakwirira kupfuura chaiyo kana chikamu chacho chiri pasi peinenge 80%, nokuti masero achiri kukwanisa kuderedza dhayi haafaniri kunge achiri kukwanisa kubereka. Zvinyorwa zvekubika doro zvinotsanangura kusavimbika kwakafanana kana chikamu chemasero mapenyu chaderera zvikuru. Tora methylene blue senzira inokurumidza yekufungidzira, uye simbisa zvisarudzo zvakakosha zvine chokuita nechikamu chidiki chemasero mapenyu, zvakadai sekushandisa zvakare mbiriso pakubika doro, nekuverenga pamapureti kana kushandisa dhayi rinopa humwe ruzivo runotsigira.

Zvinokonzerwa nemamiriro emusanganiswa ndizvo zvechitatu. Nzira yeOIV inoshandiswa pedyo nepH 4.6 uye inoyambira kuti sampuli dzine pH yakaderera uye dzinopikisa zvikuru kuchinja kwepH dzinodzivisa kushanda kwakarurama kwedhayi, ichikurudzirawo kusashandisa muedzo uyu kana shuga iri pamusoro pe100 g/L, apo masero anoonekwa aine bhuruu yakareruka inotsausa. Muedzo wekuenzanisa pH pakupenda wakawana kuchinja kukuru kwekukwanisa kuona misiyano pamazinga akaedzwa, zvinoenderana netsvakurudzo yenzira yemethylene blue inoenderana nepH. Tsvina ndeimwe njodzi yakazvimirira: tunhu twakapendwa mumisanganiswa yezviyo ine tsvina tunofanana nemasero akafa uye tunowedzera kusiyana kwezvinowanikwa nevaongorori vakasiyana.

Kusiyana uku kwakayerwa, hakusi kungotaurwa nezvakamboonekwa. Mumuedzo wemwedzi mitanhatu panzvimbo yekubika doro wakataurwa mukuenzanisa mbiriso kweFEMS, zviyero zvemethylene blue zvakaitwa kaviri paavhareji zvaiva nekutsauka kwakajairika kwe6.1%, zvichienzaniswa ne0.2% yeoxonol flow cytometry, uye vaongorori vaviri vaisiyana kusvika pamapoinzi 13 muzana pasampuli imwe chete. Kuenzanisa kwakayerwa kwenzira dzekuongorora masero mapenyu kwakaitwa muna2023 kwakawana kuti fluorescence microscopy neflow cytometry zvaipa zviyero zvakanyatsoenderana zvikuru, uku zvichiratidza kuti kushanda kwenzira kunoenderana nerudzi rwembiriso nemamiriro emusanganiswa. Tsananguro yekare yenzira yakaitwa naPainting naKirsop (PMID 24430080) ichiri hwaro hwekutarisa, uye wongororo yakafara yemiedzo yehupenyu nekushanda kwemasero embiriso (PMID 25154541) inoisa methylene blue pakati pemiedzo inotsigirana pane inoita mhedziso yekupedzisira. Trypan blue haisi iyo nzira iri nani pasina kumwe kufunga pano: inoratidza kurasikirwa nekusakuvadzwa kwemembrane, ukuwo methylene blue ichiratidza kupinda kwedhayi pamwe nebasa rekuderedza, saka masero ari pasi pekushushikana anogona kuiswa mumapoka akasiyana nemadhayi maviri aya.

Muedzo wekufema: yera nguva inotorwa kuti bhuruu ipere

Muedzo wechipiri hautomboshandisi maikorosikopu. Mbiriso pamwe neglucose pamwe nemethylene blue yakadilutwa zvinoiswa muchubhu yakavharwa, uye wachi inoyera nguva inotorwa kuti bhuruu ipere. Masero paanenge achiita metabolism, anogadzira zvinhu zvinopa maerekitironi pakuderedza, zvinoshandura dhayi rebhuruu kuti rive risina ruvara, ukuwo okisijeni yakanyunguduka ichiita okisidhesheni yoridzorera. Kana masero aderedza okisijeni, kuderedzwa kunozokunda pakupedzisira uye chubhu inoshaya ruvara. Naizvozvo nguva yekupedzisira inoratidza kuenzana pakati pekufema, kuwanda kwedhayi, nekuwanika kweokisijeni, uye inofanira kutaurwa senguva yekuderedzwa kana kupera kweruvara rwemethylene blue, kwete semwero wakananga wekushandiswa kweokisijeni yemamorekuru.

Shandisa dhayi rakadilutwa zvakanyanya kupfuura pakuyera chikamu chemasero mapenyu kuitira kuti nguva yacho ive nyore kuyera. Bhuku reAQA A level remiedzo inodiwa rinoisa izvi pamwero mumwe zvakanaka: 1 g glucose pa100 mL mvura yadziyiswa kusvika pa30 C, 5 g mbiriso yakaomeswa inowedzerwa nguva pfupi isati yashandiswa, uye dhayi rekushandisa riri pedyo ne0.001% rinogadzirwa nekudiluta 0.1 mL yemusanganiswa wekutanga we1% kusvika pa100 mL. Sanganisa 2 mL yemusanganiswa wembiriso neglucose ne2 mL yedhayi rekushandisa, rega zvienzane netembiricha yemuedzo, zunza kamwe chete kwemasekonzi 10, tanga kuyera nguva, uye usazunzazve. Kuzunza kunodzorera okisijeni uye kunoita kuti chubhu ive bhuruu zvakare, chinova ndicho chikanganiso chikuru chinokonzerwa nemaitiro. AQA inoti musanganiswa uyu unofanira kazhinji kushaya ruvara mukati memaminitsi anenge 5 pa35 C, neavhareji dzemiedzo dze275 s pa20 C, 128 s pa35 C, uye 145 s pa45 C. Iyi mienzaniso kubva pagadziriro imwe chete, kwete miyero inoshanda kwose kwose. Ukama hunoyerwa pakati pemwero wekuderedzwa nembiriso iri kuita metabolism mumamiriro akadzorwa hunotsigirwa netsvakurudzo dzakanangana nekuderedzwa, kusanganisira basa rakarongwa sePMID 25773544 netsvakurudzo yemashandiro acho iri muYeast (2014).

Muenzaniso wekuchinja nekufamba kwenguva (wakagadzirwa nokuda kwechinyorwa chino)

Tafura iri pasi apa muenzaniso wakagadzirwa nokuda kwechinyorwa chino uchishandisa gadziriro yemhando yeAQA yakatsanangurwa pamusoro: 2 mL yemusanganiswa wembiriso neglucose pamwe ne2 mL yedhayi rekushandisa riri pedyo ne0.001%, kuzunza kamwe kwemasekonzi 10, wozosiya musanganiswa uri patembiricha yakatarwa usingakanganiswi. Zvakanyorwa zviyero zvemuenzaniso zvakarongwa nenzira inofanira kunyorwa neboka revadzidzi, chiratidzo chemwero chichitsanangurwa se1 inopatsanurwa nenguva yekupera kweruvara mumasekonzi. Ita muedzo nemachubhu mamwe chete uye tsiva zvakanyorwa izvi nezviyero zvawawana.

Dhirowa girafu rechiratidzo chemwero zvichienderana netembiricha yemachubhu A, B, naC. Mutsetse unokwira wobva wakati sandara kana kudzika, unova mufambiro unotarisirwa unokonzerwa nemaenzayimu, kwete mutsetse wakatwasuka. Machubhu D, E, naF haasi mapoinzi ezviyero pamutsetse iwoyo. Ndiwo miedzo yekuenzanisa inoita kuti mutsetse ududzirike.

Miedzo yekuenzanisa inoita kuti muedzo mumwe nomumwe uve nezvaunoreva

Muedzo wose wekufema unoda mimwe mitatu inouperekedza. Mbiriso yakabikwa kana kuurayiwa pamwe neglucose nedhayi inofanira kuramba iri bhuruu, zvichiratidza kuti kuderedzwa kunokurumidza kunoenderana nemetabolism yemasero mapenyu. Mbiriso ipenyu pamwe nedhayi pasina glucose yakawedzerwa muedzo wekuenzanisa unobata chinodyiwa nemasero, asi tarisira kuti ruvara rucharamba ruchipera zvishoma nezvishoma nokuti mbiriso inoshandisa zvayakachengeta mukati. Glucose pamwe nedhayi pasina mbiriso muedzo wekuenzanisa makemikari chete, unokosha nokuti glucose iri mumamiriro earukari inogona kuderedza methylene blue pasina masero aripo.

Kuverenga masero mapenyu kunoda rumwe rudzi rwekuenzanisa: kusachinja kwemaitiro. Musanganiswa wedhayi wekutanga mumwe chete, chiyero chekusanganisa chimwe chete, nguva yekupenda imwe chete, nzvimbo yekamuri imwe chete, mitemo yemabukira neyemiganhu mimwe chete, uye kuverenga usingazivi kuti sampuli ndeipi pazvinokwanisika. Kana vaongorori vaviri vakasiyana nemapoinzi gumi, pepa rekuverengera rinowanzoratidza chikonzero: nguva dzakasiyana, muganhu wakasiyana wekuti bhuruu yakadii ndiyo inoverengwa, kana mitemo yakasiyana yemabukira. Nzira dzekugadzirira nekuona masero akapendwa dzine ukama nenzira dzakapfava dzekupenda mabhakitiriya, ukuwo ongororo yakafara yezvinoratidzwa nedhayi mumasampuli akasiyana iri muatirasi yekupenda kunoongororwa nemaikorosikopu. Pamusoro pefizikisi iri shure kwekuyera kusimba kwebhuruu nenhamba, gwaro rekunongedzera nezvekutorwa kwechiedza rine mamiriro akanyorwa rinotsanangura nei koefishenti imwe neimwe ichifanira kuperekedzwa nemamiriro echinonyungudutsa, pH, nekuwanda kwemushonga.

Sarudza muedzo zvichienderana nemubvunzo. Kuti masero mangani ari mapenyu iye zvino mubvunzo wekuverenga, unopindurwa pamaikorosikopu nepepa rekuverengera. Kuti mbiriso iyi iri kufema zvakadii mubvunzo wemwero wekuita, unopindurwa newachi yekuyera nguva nemachubhu ane mamiriro akadzorwa. Methylene blue inoshanda pazvose, asi kwete muchubhu imwe chete panguva imwe chete.