Upload
st-johns-laboratory-ltd
View
138
Download
1
Embed Size (px)
Citation preview
HOW DID OUR ANTIBODY PERFORM?
ANTIBODY CUSTOMER REVIEW: Phospho-PP1α (T320) Polyclonal Antibody (STJ91134)
What was used?Primary antibody: STJ91134 Phospho-PP1α (T320)
Provider: St John's Laboratory
Dilution ratio: 1:500
Application: Western Blot
Materials for validation: HEK cell line
Click for product data sheet PDF
What Was The Protocol?Treatment of Materials Cells were cultured to full confluence plating density. Lysis occurred with protease inhibitors
(Roche), 1% triton, 50 mM Tris-HCL, 150 mM NaCl and a phosphatase inhibitors cocktail (Sigma). 5X of sample buffer was then added.
Gel Electrophoresis 15% polyacrylamide gel, constant voltage 160 V, for 60 minutes.
Transfer 0.45 µm PVDF membrane. 1 piece of gel (semi-dry transfer), constant current 100V, for 60 minutes.
Blocking 1X TBST with 5% BSA for 1 hour.
Primary Antibody Probing Primary antibody was added to the solution of 1X TBST with 5% BSA, 4˚C and left overnight.
Membrane Wash Membrane was washed with 1X TBST 3 times.
Secondary Antibody Probing Secondary antibody was added to the solution of 1X TBST with 5% BSA with a 1:2000 dilution ratio, for 1.5 hours.
Membrane Wash Membrane was washed with 1X TBST 3 times.
Visualization Visualization: ECL visualization for 3 min (the visualization time was determined by actual result. It was ensured that the visualization time was identical for the same antigen).
Gel electrophoresis information: 15% polyacrylamide gel, constant voltage 160 V, 60 min.
Transfer information: 0.45 nm PVDF membrane, 1 piece of gel (wet transfer), constant voltage 100 V 60 min.
No. Antigen Loading amount Primary antibody
Primary antibody
dilution ratio
Secondaryantibody dilution
ratioTarget band KD Visualization time
1 Hek lysate 40ug STJ91134 phospho PP1 1:500 1:2000 ~37KD 60S
What were the results?
What did the customer think?
Antibody Specificity:
Antibody Rating:
Testing results were provided independently by S. Hilfiker at Instituto de Parasitología y Biomedicina "López - Neyra".
JOIN THE ANTIBODY VALIDATION PROJECT
1. Order any STJ9 catalogued product using this online form.
2. Test it in your laboratory using your preferred method.
3. Return a fully completed review form to receive rewards.
…in 3 simple steps…
@StJohnsLabs
http://www.slideshare.net/stjohnslabs