Gram staining

Gram staining is a staining to differentiate between gram positive and gram negative bacteria.
Procedure
Take the slide and clean it with 70% ethanol
Take the inoculum from the sample
Spread it on the slide with the help of inoculation loop
Now add crystal violet drop by drop
Leave it for one minute
Rinse with the water
Add iodide dye
Then wash with 70 %ethanol drop by drop
Now add safranine
Observe under the microscope
Gram negative will be pink and gram positive will be purple

Use of crystal violet : gram pain bacteria having a more quantity of lipopolysaccharide so crystal violet attach to lipopolysaccharide and gives violet colour
Use of iodide : iodide attaches with crystal violet and make strong bond
Use of 70% ethanol : ethanol removes the excessive dye as gram negative bacteria do not have that much amount of lipopolysaccharide so all the remaining crystal violet and iodide washes away with the 70% ethanol
Use of safranine : safranine reacts with the lip polysaccharide of gram negative and gives pink colour