BACTERIAL SMOKING TECHNIQUE Bacteria or other microbes can be seen with a light microscope without or with staining. Bacterial staining generally uses more than one painting level. The painting results are influenced by several factors, such as: fixation, substrate, decolorizer and so on. In making bacterial smears that are ready to be stained, fixation needs to be carried out first, which aims to include: a). prevents the shrinking of cell protein globules, b). change paint affinity, c). prevent cell autolysis, d). can kill microbes quickly without causing changes in their shape or structure, e). attach bacteria to the glass slide and f). makes cells stronger/harder. The fixation method is to make a layer of bacterial suspension/smear on a glass object, then dry it in the air and pass it several times over the flame of a spirit lamp. TOOLS AND MATERIALS - Sterile petri dishes, sterile test tubes, test tube rack, volume pipette, measuring cupsterile, round and straight Oose needle, glass slide - Hand wipes, tissue, tripod, asbestos gauze, Bunsen, Incubator 37°C - Nutrient Agar Media, Nutrient Broth Media, and sterile NaCl - Bacterial culture: Staphylococcus aureus, Bacillus subtilis, Escherichia coli, and Pseudomonas aeruginosa EXPERIMENT PROCEDURE ASEPTIC TECHNIQUE 1. Adjust the Bunsen flame so that a blue flame is obtained to obtain a “sterile” area. Let the fire burn for approximately 10 minutes before working. 2. Flamb the tweezers, the cotton covering the test tube to be used is pulled slowly with tweezers in such a way that it is enough to be pulled by pinching it between the little finger and the palm of the right hand. The test tube was placed in the tube rack to the left of the practitioner. 3. The Ose needle is ignited until the tip glows, then the heating is continued slowly towards the needle handle until the wire limits. This work was carried out three times. 4. With the left hand, take the test tube containing the inocula. The cotton covering the tube is pulled by pinching it between the little finger and the palm of the right hand. The open mouth of the tube was flammable quickly. 5. The Ose needle is inserted into the inocula test tube without touching the wall to collect the inocula. Then the Ose needle is pulled out and held at a distance of 10 cm around the Bunsen flame. 6. The mouth of the test tube is flammable, then the cotton cap is put back into the mouth of the tube. The tube is returned to its shelf. 7. The tube containing the sterile medium to be inoculated is taken with the left hand. The cotton plug is pulled with the right hand, by pinching it between the little finger and the palm of the hand. Meanwhile, the Ose needle containing the inocula is still held in the right hand. The mouth of the tube is flammable. 8. The Ose needle is inserted into a tube containing sterile media and inoculation (scratching or puncture) is carried out. The Ose needle is pulled out, the mouth of the tube is flammable then the cotton stopper is reinserted and then the tube is placed back in the rack. 9. The Ose needle is flammable until it glows three times, before being stored againrack. 10. Put a label/label on each tube containing the culture.