Application
Western blot
Sample
African Green Monkey Cell lysate - whole cell (Vero cells)
Loading amount
10 µg
Specification
Vero cells
Treatment
Infected with HSV1 (KOS strain)
Gel Running Conditions
Reduced Denaturing
Blocking step
Milk as blocking agent for 30 minute(s) · Concentration: 5% · Temperature: 20°C
Other product details
Dilution
1/1000
Incubation time
16 hour(s) and 0 minute(s) · Temperature: 4°C · Diluent: blocking buffer (PBS Tween Milk)
Secondary antibody
Name
Non-Abcam antibody was used: Anti Mouse Amersham
Host species: Donkey
Clonality: Polyclonal
Conjugation: Horse Radish Peroxidase
Host species: Donkey
Clonality: Polyclonal
Conjugation: Horse Radish Peroxidase
Dilution
1/3000
Detection
Detection method
ECL Pierce
Exposure
30 second(s)
Bands
Specific: Just under 97 kDa Non-specific: None kDa
Negative control
Mock infected cells
Additional data
Additional Notes
Vero cells were mock (lane 3) or HSV1 (strain KOS) (lane 4) infected for 16 hours (MOI of 2). Human HeLa cells were also analysed (lane 1: Mock infected, lane 2: HSV1 KOS infected). Whole cell lysates were prepared (lysis in 0.5% NP40 for 15 minutes on ice, then clarification by a 10 minutes centrifugation at 14K rpm in an eppendorf centrifuge) and 10 microgrammes of the mock infected and KOS infected lysates were loaded on the 4-12 % NuPage gel. Gels were run in MOPS-SDS buffer. Although the apparent size of the protein detected seems to be a bit low for gC (calculated mmw is around 120 kDa), its is probably due to the NuPage gel used to run the samples. Please also note that a lower sized faint band also appears after a longer ECL exposure, probably representing the unglycosylated form of gC.
The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.
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提交于 Sep 05 2007