The customer used the VAPB C-term polyclonal antibody (STJ47066) from St. John's Laboratory in a Western blot with A549 cell lysates. They detected a target band at 30 kDa and observed possible dimerization or post-translational modifications of VAPB. The customer provided positive feedback that the antibody detected the target band in the cell line as expected. St. John's Laboratory invites customers to test their products for free and provide reviews to receive discounts on future orders.

1. HOW DID OUR
ANTIBODY PERFORM?
CUSTOMER REVIEW: VAPB C-term Polyclonal Antibody (STJ47066)

2. WHAT WAS USED?
Primary antibody: STJ47066 VAPB C-term Polyclonal
Antibody
Provider: St John's Laboratory
Dilution ratio: 1:800
Application: Western Blot.
Materials for validation: A459 cell lineFigure 1. Western blot analysis of VAPB
in asynchronous (lane 1) and quiescent
(lane 2) A549 whole cell lysates.

3. WHAT WAS THE PROTOCOL?
Treatment of materials: Cells were cultured to 70%-80% plating density, rinsed
with PBS, lysed with standard lysis buffer, boiled for 5min, and passed through
needle to disperse any clumps. Lysates were resuspended using 4x Laemmli buffer.
Gel electrophoresis: 12.5% polyacrylamide gel, constant voltage 160 V, 60 min.
Transfer: Nitrocellulose membrane, whole gel (wet transfer), constant voltage 100
mA overnight.
Blocking: 5% milk in 1X TBST, 1 hr at room temperature.
Primary antibody probing: Primary antibody was added to the solution of 1X
TBST with 5% skimmed milk powder, 4˚C for overnight.
Membrane wash: 1X TBST wash, 3x 5 min. each.
Secondary antibody probing: Secondary antibody (anti-rabbit IgG) added at
1:500 in 1:1 2%BSA:5% milk for 1 hour at room temperature.
Membrane wash: 1X TBST, 3x 5 min. each.
Visualization: ECL, 2 min exposure.

4. WHAT WAS THE PROTOCOL?
Gel electrophoresis information：12.5% 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
Secondary
antibody
dilution ratio
Target band
Visualization
time
1 A549 lysate 20 µg
STJ47066.
VAPB C-term
Polyclonal
Antibody
1:800 1:500 30 KDa 2 mins

5. WHAT DID THE CUSTOMER THINK?
Testing results were provided by Meagan Donaldson from
Albert Einstein College of Medicine. They stated that “The
target band was found in the cell line. Upper band may
correspond with reported dimerization of VAPB with other
VAMP family members or posttranslational modifications.
NOTE: samples were NOT boiled before loading onto gel.”

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with the best product, but to also give them
the opportunity to test the product’s
performance risk free.

7. Contact Us: info@stjohnslabs.com
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