1. Technical Support Questionnaire – Western Blot
Name: Ching-An Wu
Catalog #: NB100-105
Lot Number: AD-1.
PO/Order Number: 20120913004
Species, Tissues or Cell Lines Tested: HeLa cell lysate with hypoxic treatment.
Test Sample Preparation: Hypoxic treatment was performed by incubating with 0.7% oxygen for 4
hours. Right after the treatment, cells were collected and placed on ice, washed with ice cold PBS,
and lysed with ice cold RIPA buffer, and then centrifuged and boiled with sample loading buffer.
Antibody Storage Conditions: 4℃
℃
Gel/Electrophoresis Conditions: 8% SDS-PAGE
Transfer Conditions: semi-dry
Blocking Solution & Duration: 5% milk in TBST.
Primary Antibody Diluent and Dilutions Tested: 1:1000 with TBST
Primary Antibody Incubation Time and Temperature: 4℃ over night
℃
Wash Solution Composition, Repetitions & Times: Click here to enter text.
Secondary Antibody Manufacturer, Host Species, Dilution, & Diluent: anti-mouse HRP at 1:5000 with

2. 5% milk in TBST
Secondary Antibody Incubation Time & Temperature: Room temp. 1hr
Wash Solution Composition, Repetitions, & Times: Click here to enter text.
Detection System, Procedure & Development Time: ECL
Molecular weight of band(s): Click here to enter text.
Controls: Click here to enter text.
Observations: Incorrect band and too many background band.