Hoʻokolohua Hemoglobin Electrophoresis

Kumu Hoao

ʻO ka Hemoglobin electrophoresis ka manaʻo e ʻike a hōʻoia i nā hemoglobin maʻamau a maʻamau.

Ma muli o nā uku like ʻole a me nā helu isoelectric o nā ʻano hemoglobin like ʻole, i loko o kahi hopena buffer pH, i ka wā i haʻahaʻa ai ka helu isoelectric o ka hemoglobin ma mua o ka pH o ka solution buffer, lawe ka hemoglobin i kahi uku maikaʻi ʻole a neʻe i ka anode i ka wā electrophoresis. ʻO ka mea like ʻole, neʻe ka hemoglobin me ka uku maikaʻi i ka cathode.

Ma lalo o kekahi volta a ma hope o ka manawa electrophoresis kiko'ī, hōʻike nā hemoglobin me nā koina like ʻole a me nā paona molekala i nā kuhikuhi neʻe a me nā wikiwiki. ʻAe kēia i ka hoʻokaʻawale ʻana o nā ʻāpana ʻokoʻa, a hiki ke hana ʻia ka nānā ʻana i ka colorimetric a i ʻole electrophoretic scanning ma kēia mau ʻāpana e helu i nā hemoglobin like ʻole. ʻO ke ala maʻamau ka pH 8.6 cellulose acetate membrane electrophoresis.

I loko o ka cytoplasm, nā pūʻulu ethylene glycol (CHOH-CHOH) i loko o nā mea glycogen a i ʻole polysaccharide (e like me mucopolysaccharides, mucoproteins, glycoproteins, glycolipids, etc.) ua hoʻoneʻe ʻia e ka waikona periodic a hoʻololi ʻia i mau pūʻulu aldehyde (CHO-CHO). Hoʻohui kēia mau pūʻulu aldehyde me ka Schiff reagent ʻulaʻula ʻulaʻula ʻulaʻula ʻole, e hana ana i kahi waihoʻoluʻu poni-ʻulaʻula e waiho i kahi polysaccharides i loko o ke keena. ʻIke ʻia kēia pane ʻana ʻo ia ʻo periodic acid-Schiff (PAS) staining, i kapa mua ʻia he glycogen staining.

ʻAno hoʻokolohua

Mea Hana:Cellulose acetatemembrane, mea hana electrophoresis(DYCP-38C a me ka lako mana DYY-6C ), Mea Hoʻouka Laʻana Kiʻekiʻe (pipette), spectrophotometer, cuvettes colorimetric, nā pale.

Hoʻopaʻa:

(1) pH 8.6 TEB Buffer: Kauona 10.29 g Tris, 0.6 g EDTA, 3.2 g boric acid, a hoʻohui i ka wai distilled i 1000 ml.

(2) Borate Buffer: E kaupaona i 6.87 g borax a me 5.56 g boric acid, a hoʻohui i ka wai i hoʻoheheʻe ʻia i 1000 ml.

Kaʻina hana:

Phoʻoponopono i ka Hemoglobin Solution

E lawe i 3 ml o ke koko me ka heparin a i ʻole ka sodium citrate ma ke ʻano he anticoagulant. Centrifuge ma 2000 rpm no 10 mau minuke a hoʻolei i ka plasma. Holoi i ke koko ʻulaʻula ʻekolu manawa me ka physiological saline (750 rpm, 5 minuke centrifugation i kēlā me kēia manawa). Centrifuge ma 2200 rpm no 10 mau minuke a hoʻolei i ka supernatant. E hoʻohui i ka nui like o ka wai i hoʻoheheʻe ʻia, a laila e hoʻohui i 0.5 mau manawa o ka nui o ka carbon tetrachloride. E lulu ikaika no 5 mau minuke, a laila centrifuge ma 2200 rpm no 10 mau minuke e hōʻiliʻili i ka hopena Hb luna no ka hoʻohana ʻana ma hope.

ʻO ka hoʻoulu ʻana i ka Membrane

E ʻokiʻoki i ka membrane cellulose acetate i loko o nā ʻāpana e ana ana he 3 knm × 8 knm. E hoʻoinu iā lākou i loko o ka pH 8.6 TEB buffer a piha pono, a laila wehe a holoi maloʻo me ka pepa kānana.

ʻike maka

E hoʻohana i ka pipette e ʻike pono i ka 10 μl o ka solution hemoglobin ma luna o ka membrane cellulose acetate (ʻo ka ʻaoʻao ʻala), ma kahi o 1.5 knm mai ka lihi.

Electrophoresis

E ninini i ka wai hoʻoheheʻe borate i loko o ke keʻena electrophoresis. E kau i ka membrane cellulose acetate me ka ʻaoʻao kiko ma ka hopena cathode o ke keʻena. E holo ma 200 V no 30 mau minuke.

Elution

E ʻoki i nā ʻāpana HbA a me HbA2, e hoʻokomo iā lākou i loko o nā paipu hoʻāʻo ʻokoʻa, a hoʻohui i 15 ml a me 3 ml o ka wai i hoʻomaʻemaʻe ʻia. E lulu mālie e hoʻokaʻawale i ka hemoglobin, a laila hui.

kala kala

Hoʻopau i ka absorbance me ka hoʻohana ʻana i ka wai distilled no ka elution solution a ana i ka absorbance ma 415 nm.

Heluhelu

HbA2(%) = Absorbance o HbA2 paipu / (Absorbance o HbA paipu × 5 + Absorbance o HbA2 paipu) × 100%

Ka helu ʻana i nā hopena hoʻokolohua

Ka Papa Kuhikuhi no ka pH 8.6 TEB Buffer Cellulose Acetate Electrophoresis: HbA > 95%, HbA2 1%-3.1%

Nā memo

ʻAʻole pono e lōʻihi ka manawa Electrophoresis. ʻAʻole pono e maloʻo ka membrane cellulose acetate i ka wā electrophoresis. Hoʻopau i ka electrophoresis ke hoʻokaʻawale ʻia ʻo HbA a me HbA2. Hiki i ka electrophoresis lōʻihi ke hoʻopuehu i ka band a me ka blurring.

E hōʻalo i ka hoʻohana nui ʻana i ka laʻana. ʻO ka nui o ka wai hemoglobin hiki ke alakaʻi i ka hoʻokaʻawale ʻana o ka band a i ʻole ka lawa ʻole o ka ʻili ʻana, e hopena i nā pae HbA wahaheʻe.

E pale i ka haumia o ka membrane cellulose acetate me nā protein.

ʻAʻole pono ke kiʻekiʻe o ke au; Inā ʻaʻole, ʻaʻole hiki ke hoʻokaʻawale nā ​​kaula hemoglobin.

E hoʻokomo mau i nā hōʻailona mai nā kānaka maʻamau a me nā hemoglobin kūponoʻole i ʻike ʻia e like me nā mana.

Hana ʻo Beijing Liuyi Biotechnology i ka pahu electrophoresis ʻoihana no ka hemoglobin electrophoresis ʻo ia ke kumu hoʻohālike.DYCP-38Ccellulose acetate membrane electrophoresis tank, a aia ʻelua mau hiʻohiʻona o ka mana electrophoresis i loaʻa no ka cellulose acetate membrane electrophoresis tankDYY-2CaDYY-6Clako ikehu.

I kēia manawa, hāʻawi ʻo Beijing Liuyi Biotechnology i ka membrane cellulose acetate no nā mea kūʻai aku, a hiki ke hoʻololi i ka nui o ka cellulose acetate membrane. Welina mai e nīnau iā mākou no nā laʻana a me ka ʻike hou aku.

Aia ʻo Beijing Liuyi brand ma mua o 50 mau makahiki moʻolelo ma Kina a hiki i ka hui ke hāʻawi i nā huahana kūpaʻa a kiʻekiʻe a puni ka honua. Ma o nā makahiki o ka hoʻomohala ʻana, kūpono ia i kāu koho!

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