Negative Staining of Phage Samples

For negative staining, 5 µL of PEG purified phage sample (~ 6 × 10⁶ pfu/ml) was applied onto glow-discharged carbon coated grids and incubated for five minutes at room temperature. Grids were washed with 5 µL of deionized water before incubating for 2 min with a 1% uranyl acetate solution. Electron micrographs were taken using a Phillips CM-10 microscope at the Manawatu Microscopy and Imaging Centre (MMIC, Massey University, Palmerston North, New Zealand)^{55 (link)}.

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Sattar S., Ullah I., Khanum S., Bailie M., Shamsi B., Ahmed I., Shah S.T., Javed S., Ghafoor A., Pervaiz A., Sohail F., Shah N.A., Imdad K., Bostan N, & Altermann E. (2022). Phenotypic characterization and genome analysis of a novel Salmonella Typhimurium phage having unique tail fiber genes. Scientific Reports, 12, 5732.

Publication 2022

CM-10 microscope

Carbon Electron Microscopy Phage Uranyl acetate

Corresponding Organization : COMSATS University Islamabad

Other organizations : AgResearch, University of Veterinary and Animal Sciences, Massey University, Riddet Institute

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Variable analysis

independent variables

Volume of PEG purified phage sample applied onto grids (5 µL)
Concentration of phage sample (~ 6 × 10^6 pfu/ml)
Duration of incubation on grids (5 minutes)
Concentration of uranyl acetate solution (1%)
Duration of incubation with uranyl acetate solution (2 minutes)

dependent variables

Electron micrographs of the phage samples

control variables

Room temperature for incubation
Use of glow-discharged carbon coated grids
Washing grids with deionized water

controls

Positive control: Not specified
Negative control: Not specified

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