Premix WST-1을 이용한 세포 생존능력 측정 Premix …

1 14 Life Science & Biotechnology Premix WST-1 Premix WST-1 Cell Proliferation Assay System Technical Tip 2 Premix WST-1 Cell Proliferation Assay System Tetrazolium Salts (WST-1) formazan ELISA . (Dehydrogenase) succinate-tetrazolium Reductase ( ) . Premix Premix WST-1 Cell Proliferation Assay System (TaKaRa Code MK400).

2 (2,500 ) Premix WST-1 25 ml x 1 bottle * 100 l/well 10 l Premix WST-1 **100 l/well 2,500 ** Premix WST-1 WST-1 electron coupling Phosphate buffered saline (PBS) . 1. Microtiterplate 100 l/well .** . ~5 104 /well, 24~96 . 2. , Premix WST-1 1 well 10 l .3. 1 4 . 4. ELISA reader background *.

3 Formazan ELISA 420-480 nm ( 440 nm)( 2), 600 nm .* Background : 1 100 l Premix WST-1 10 l , ELISA reader background ( + Premix WST-1 ) . Premix WST-1 , (incubation) , . 2 background . Premix WST-1 (incubation) ( ) Premix WST-1 , ( , , 1, 2, 4 ).

4 Premix WST-1 . Premix WST-1 Cell Proliferation Assay System WST-1 formazan . 1 : T CTLL-2 Inter-leukin 2 (IL-2) 10 % heat-inactivated fetal calf serum (FCS), 2 mM L-glutamine, 1 mM Na-pyruvate, 1 , 50 mM 2-mercaptoethanol RPMI 1640 ( , , ) IL-2 (10,000 U/ml, 5 g/ml, ) Premix WST-1 1. Microtiter plate IL-2 ( , ng/ml) 100 l , CTLL-2 4 103 cells/well seeding.

5 2. 37 C, 5% CO2 48 .3. Premix WST-1 well 10 l , 37 C, 5 % CO2 4 . 4.. 2 : WEHI-164 TNF- 1. Tetrazolium salt (WST-1) formazan (EC=electron coupling reagent, RS=mitochondrial succinate-tetrazolium-reductase system)September 2009 15 Technical Tip 10 % heat-inactivated fetal calf serum (FCS), 2 mM L-glutamine, 1 g/ml actinomycin D RPMI 1640 ( , , ) TNF- (10 g/ml, ) Premix WST-1 1. 1 g/ml actinomycin C1 1 x 106 cells/ml WEHI-164 37 C, 5 % CO2 3.

6 2. Actinomycin C1 (1 g/ml) 100 l 5 x 104 cells/well , TNF- ( ng/ml) .3. 37 C, 5 % CO2 24 .3. Premix WST-1 well 10 l , 37 C, 5 % CO2 4 , . 4.. Q & AQ1: MTT, XTT, MTS (tetrazolium salt) Premix WST-1 Cell Proliferation Assay System ? A1: , tetrazolium salt formazan . MTT formazan , . , XTT MTS formazan Premix WST-1.

7 Tetrazolium salt WST-1 , . Q2: , Premix WST-1 ?A2: ( , ), , overnight (16 hr) .Q3: Formazan plate reader ? A3: 420~480 nm . , 440 nm , . 3. IL-2 CTLL-2 4. WEHI-164 TNF- TaKaRa CodeLDH Cytotoxicity Detection Kit2000 testsMK401in situ Apoptosis Detection Kit20 MK500 PBS (Phosphate Buffered Saline) Tablets200 T900 2.

8 Premix WST-1 Cell Proliferation Assey System <A> (formazan) <B>. Premix WST-1 Cell Proliferation Assay System 10% FCS RPMI1640 (BioWittaker ) 1:10 .absorbancewavelength (nm) 450 500 550 600 650 700