Visualizing the Effect of pH on Solubilization of the Influenza A Viral Core

Begin with purified influenza A viral core fractions, treated with various pH conditions, ranging from neutral to acidic.
The viral core fraction contains matrix proteins and nucleoproteins.
Heat the samples to denature the proteins.
Load onto a polyacrylamide gradient gel.
Initiate electrophoresis to separate the proteins based on molecular weight, forming distinct bands.
Incubate the gel in a fixation solution to immobilize the proteins.
Wash to remove fixative, add colloidal Coomassie dye, and incubate. The dye binds to proteins, allowing visualization of the bands.
Remove the dye and wash to destain the gel, which enhances band visibility.
Capture a high-resolution image of the gel.
At neutral pH, the higher intensity of the matrix protein and nucleoprotein bands indicates that the viral core remains intact and retains associated proteins.
As pH decreases, band intensity progressively diminishes.
This indicates that acidic conditions trigger viral core disassembly and promote protein solubilization during purification.

To carry out SDS-PAGE, heat all the samples at 95 degrees Celsius for 10 minutes. Load 20 microliters of the dissolved pellets onto a precast gradient Bis-Tris mini-gel and run at 200 Volts for one hour in 1X MOPS SDS running buffer.

Incubate the gel in fixation solution for one hour. Rinse one to two times with water, and stain overnight in a 15-centimeter cell culture dish with a sufficient volume of colloidal Coomassie solution while gently shaking at room temperature.

Destain the gel in doubly-distilled water, replacing the liquid every 15 to 20 minutes until the gel background becomes clear. Store the gel in doubly-distilled water at four degrees Celsius until it is scanned for band quantification.