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Solution Informationhelp
Enzyme: streptokinase A precursor
inhibitor: BDBM57624
substrate: n/a
 
Solution Type: Aqueous
pH at Preparation: n/a
Temp. Prep.: n/a
Comments: Day 1 Streak out UMAA 2616 for colonies on THY/S (THY with streptomycin 100 μg/mL) plate; 37'C O/N Day 2 Grow an overnight culture in THY/S from a single colony, 37oC O/N Day 3 In the morning, dilute 1:20 of the O/N culture into fresh THY/S in flask. Monitor OD600 until it reaches 0.6-0.8 (between 3-5 hrs). Dilute the 0.8OD Culture down to OD 0.038 into cold (4'C) THY/S; stir at 4'C O/N Day 4 Dispense assay plates (Corning 3570) at 30ul/well of THY/S with Combi (Thermo) Pin Compounds (100 nL) using Cybi-Well (CyBio) Seed cells (warmed up to RT) onto the pinned plates at 20ul/well with Combi 37'C for 6 hrs in Liconic incubator Pellet cells at 3000 rpm Transfer 2 x 10 uL/well supernatant to clear plates (Nunc 242757) for assay with Cybi-Well Vario (CyBio), store at -20'C. Cool the culture plates to RT for 30 minutes Add 50 uL/well of 1/2x BacTiterGlo (Promega G8233) RT 10 minutes, Read for luminescence SK activity is measured as follows:
 
 

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Last update November 1, 2007
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