E-portfolio for teknik asas mikrobiologi: E-portfolio week 8

This week I have my hands on test on streak plate and simple staining. Then, I did an experiment on capsule stain and spore stain. For spore stain, the procedures for the experiment were as followed. Individual smears were made in usual manner by using aseptic technique on two clean glass slides. The smears were allowed to air-dry and heat fix in the usual manner. Then. the smears were flooded with malachite green and placed on top of a beaker of water sitting on a warm hot-plate, allowing the preparation to steam 2 to 3 minutes. The slides were then removed from the hot plate, cooled and washed under running tap water. Next, the slides were counterstained with safranin for 30 seconds and washed with tap water. Finally, the smears were blotted dry with bibulous paper and examined under oil immersion. The organisms used are Bacillus cereus and Clostridium sporogenes.

For capsule stain, I did the following procedures.The procedures of the experiment were as followed. One glass slides was obtained and several drops of crystal violet stain were placed on the slide. By using aseptic technique, three loopfuls of Klebsiella pneumonia were added to the stain and mixed gently with the inoculating loop. Then with a clean slide, the mixture was spreaded over the entire surface of the slide to create a very thin smear and the slide was let stand for 5 to 7 minutes. The smear was allowed to air-dry. The smears were washed with 20% copper sulphate solution. Finally, the smears were blotted dry and examine under oil immersion. Step 1 to 5 was repeated for Enterobacter aerogenos.

The procedures of negative staining were also done to observe the capsule of the organisms by using nigrosin. The procedures for negative stain were as followed. First, a small drop of nigrosin was placed close to one end of a clean slide. Then, a loopful of inoculum from the Klebsiella pneumonia culture was placed in the drop of nigrosin by using aseptic technique and mixed. Next, a slide was placed against the drop of suspended organisms at 45° angle and the drop was allowed to spread along the edge of the applied slide. The slide was then pushed away from the drop of suspended organisms to form a thin smear. Steps 1 to 4 were repeated for slide preparation of the remaining cultures. The slides were then examined under oil immersion.

Slides of the microorganisms for spore stain,capsule stain and negative stain