A student from one of the research labs is having trouble preparing a slide for examination and photographing. The bacterial slide that he has brought to you was prepared using a commercially purchased stain. He has asked for your help in determining what he is doing wrong so that he can change the lab protocols and continue on with his project. After examining the slide under oil immersion, you determine that no bacteria are present even though the student is able to show you the culture he used to make that slide that has visible growth in the liquid medium.
Which of the following statements does NOT explain the fact that there are no bacteria present on the student’s slide?

a. By not allowing a glass slide to completely air dry before heat fixation, the flame will cause the surrounding water to boil and this will damage the bacterial cell.
b. Overheating during the fixation step boiled the water within the bacterial cells and resulted in the cells bursting.
c. Insufficient heating of the slide did not drive out the thin layer of water and this resulted in minimal bonding between the bacteria and the glass slide.
d. Rinsing with alcohol during the washing step stripped the bacteria off the glass slide.

Alcohol is used to fix the slide means alcohol help the bacteria's cell to fix with slide or attach with slide so this is the wrong explanation. And mostly, washing is done with running tap water.

Other statements can be true for missing of bacteria on slide like incomplete air dry results in damage of bacterial cells from heat fixation. Overheating will also burst the cell by boiling of water present in bacterial cell. Insufficient heat will not completely fix the bacterial smear to the slide and due to minimal bonding the bacteria will wash out from the slide during washing step.

So, alcohol is use to fix the bacterial smear to slide and does not use to remove bacteria from slide.