Hi,
I am trying to use lateral flow assay to detect the whole intact virus by targeting the envelop proteins. That is, I use antibodies against the envelop protein of the virus for my assay. I conjugate this Ab to Gold NP as the label.
I am trying both Sandwich and Competitive Lateral flow assay. Tried with several Ab but without any success.
Recently, purchased commercially available Ab raised against the envelop protein. I even bought the envelop protein against which the antibodies are raised. The assay worked with these set of Abs and the commercial envelop protein.
But, when I tried repeating the assay with the same Abs against the whole virus cultured in our lab, we are not getting the desired result.
What could be the reason that the assay is working with the envelop protein (commercial one) of the Virus but not with the Whole Virus ?
Is it that the envelop protein of the whole virus is not having its epitopes properly exposed ?
Is there a way I can breakdown the whole virus by some treatment so that envelop protein becomes easily accessible ?
Would really appreciate your valuable suggestions
Thank You for your time
Try heat treatment.
Add detergent to the virus sample.
Did you check at ELISA thatcantibodies react with antigen.
Hi Daniel,
We are working on Zika Virus. Are CD 63 & CD9 proteins are expressed on the Virus envelop (if the virus was cultivated in human cells). Are these proteins easily targeted ?
Hi Oded Babai,
Yes, I checked. The antibodies react with the antigen (recombinant envelop protein). But the antibodies are not detecting the whole intact virus on a lateral flow set up.
I used 1000 P.F.U. and 100000 P.F.U of virus for the study
Thank you very much for your response
If the antibody raisd against specific protein or the whole virus it can explain the result. Can you check the growth medium for secreted antigen?
You can do western blot to see which protein ofvthe virus react with your antibodies.
Hi Oded Babai,
The antibody is raised against specific part of envelop protein (400 aminoaids) and not the whole virus.
Doing a western is a good idea. Thank you very much
Hi,I have two questions
How antibodies and conjugated antibodies located on the nitrocellulose membrane in lateral flow test?
How sample , adsorbents, conjugates pads and nitrocellulose membranes need pretreatment in lateral flow test?
Thank You for your time
Hi
I suggest that before using the strip test, you first optimize the reaction conditions with the ELISA method.
I work in the field of bacterial diagnosis. I hope the following files are effective for you.
1. Optimisation of immuno-gold nanoparticle complexes for antigen detection
2. Quantifying Bound and Active Antibodies Conjugated to Gold
Nanoparticles: A Comprehensive and Robust Approach To Evaluate
Immobilization Chemistry
3. Development of new lateral-flow immunochromatographic strip using colloidal gold and mesoporous silica nanoparticles for rapid diagnosis of active schistosomiasis