Article New low-viscosity overlay medium for viral plaque assays

This paper should be very helpful for you. Use of Avicel will be better for plaquing flu.

Thank you so much for your kind response. I will try it.

I think the thing you are looking for is the protocol of ''plaque assay''

1. Day 0

Seed cells in 6 well within 10 %FBS.

2. Day 1

1/ Discard the medium, wash cells with PBS one time

2/ Add virus (10X serial dilutions from the prior well for each well)

3/ Incubate 37'C for 1 hour, pat the 6 well dish every 15 minutes should

be helpful to the homogeneous infection.

4/ Discard the medium containing virus.

5/ Wash cell with PBS three times.

6/ Heat the agarose and mix with medium

7/ Add the mixture to the cell.

8/ Wait around 15 minutes. After the agarose turn into solid phase, put the

6 well dish into 37'C incubator.

[Notes]

* As treating influenza virus, the medium should contain trypsin to help

virus infection.

* As treating influenza virus, the medium should not contain FBS since the

antibodies within FBS may attack influenza virus.

* Be careful that step 7 is a critical step. The mixture should be around 37

'C or the agarose can get solid. Utilizing 37'C water bath may help you

maintain the temperature.

3. Day 3 (or the time you yearn to terminate virus infection)

Fix cells with formaldehyde.

4. Day 4

1/ Discard formaldehyde.

2/ Use water flow to remove agarose.

3/ Use crystal violet to visualize the viral plaques.

Wish it would be helpful

Wow, dear Lou Tzu Yu thank you so much for this exaustive protocol. I have set up in lab similar one, thank you so much, now I can compare them and see where do I stand.

I wish you a nice day,

Regards,

Brankica