HOW DID OUR ANTIBODY PERFORM?

ANTIBODY CUSTOMER REVIEW: Caspase-1 Polyclonal Antibody (STJ22893)

What was used?Primary antibody: CASP1 Polyclonal antibody (STJ22893)

Provider: St John's Laboratory

Dilution ratio: 1:1000

Application: Western Blot

Materials for validation: HCT116 and U937 cells

Click for product data sheet PDF

What was the protocol?Treatment of Materials Cells were lysed in 1.5 mls of 10X standard RIPA lysis buffer with protease

inhibitors (PMSF and aprotinin) added. After lysis, samples were spun down and 4X sample buffer added. About 35 ul of the lysed cells was separated by SDS-PAGE at 12.5% and then transferred and blotted

Gel Electrophoresis 12.5% polyacrylamide gel, constant voltage 1200V, 60 min. 10 ug per lane.

Transfer Nitrocellulose membrane

Blocking 1X TBST with 10% skimmed milk powder for 45 minutes.

Primary Antibody Application 1:1000 primary antibody was added to the solution of 1X TBST with 2.5% skimmed milk powder, 4˚C for overnight.

Secondary Antibody Application Secondary antibody (anti-rabbit HRP) was added to the solution of 1X TBST with 2.5% skimmed milk powder with 1:7500 dilution ratio, for 1.5 hours.

Visualisation ECL kit

What was the protocol?Gel electrophoresis information, Gel 1 and 2： 12.5 % polyacrylamide gel, constant voltage 120 V, 60 min.Transfer information： Nitrocellulose, 1 piece of gel (wet transfer)

Gel No. Antigen Loading amount Primary antibody Primary antibody dilution ratio

Secondaryantibody dilution ratio Target band kDa Visualization time

1 HCT116, No stimulation 10 ug STJ31393Caspase 3 or 1 1:1000

1:7500 See below See above

2 HCT116, LPS/ATP 10 ug

3 HCT116. TNFa/CHX 10 ug

4 HCT116, Doxirubicin 10 ug

5 HCT116, staurosporine 10 ug

6 U937, No stimulation 10 ug

7 U937, LPS/ATP 10 ug

8 U937, TNFa/CHX 10 ug

9 U937, Doxirubicin 10 ug

10 U937, staurosporine 10 ug

What were the results?

(St John’s Laboratory)

“Caspase 1 antibody recognizes intact and cleaved products in HCT116 human colon cancer cells and in U937 human monocyte lymphoma cells.”

Testing results were provided independently by Professor Shairaz Baksh at the University of Alberta. They stated that:

What did the customer think?

Antibody Specificity:

Antibody Rating:

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