ASEPTIC TECHNIQUE

ASEPTIC TECHNIQUE Introduction The purpose of the Aseptic technique in the lab is to prevent the patient from getting an infection and to prevent the spread of pathogens. The goal of aseptic technique is asepsis, which means the absence of pathogenic organisms in the clinical setting. We are investigating the proper techniques to keep contamination to a minimum and make sure we can keep a sterile environment in the lab. By following the procedures we were able to keep the specimens from getting contaminated and we were able to keep our wetlab sterile. Procedure Conduct Experiments: Experiment 1: Understanding Aseptic Technique Conduct Contamination Test -Click the NEW UNKNOWN button. Enter Label and select a subgroup. -Type TECHNIQUE1 in the “Enter a Label” line -From the Subgroup dropdown menu, Select ASCEPTIC TECHNIQUE. Do not click Auto-inoculation allowed. -Record details of your case study scenario. -Record Gram reaction by clicking appropriate radio button. -Indicate the results of the Gram Stain in the Lab Report. -From the Media dropdown, select Phenol Red Glucose Broth Durham Tube. -Enter PhenolRedGlucose in the Medium Label window to label your sample. Two tubes will appear on the workbench. Take a careful look at the sample tube. Is there media in the Durham Tube? -Note the status of the Traffic Lights for Inoculation and Contamination in the upper right hand corner of the lab software window. -Right click on the Phenol Red Tube and select REMOVE CAPS/LIDS. -What color are the Inoculation/Contamination traffic lights? -Type TECHNIQUE1 in the “Enter a Label” line -From the Subgroup dropdown menu, Select ASCEPTIC TECHNIQUE. -Check traffic signals to confirm you successfully inoculated your sample and no contamination was introduced. -Repeat these steps until you have successfully inoculated your medium without contamination. How many times did it take you to accomplish this? Easier or harder? -Incubate the successful inoculation for 24 hours. -Flame your loop to sterilize it and turn off the Bunsen Burner -Select POINTER from Tool Menu -Point and drag the inoculated sample to the 37 degree incubator -Click the NEW DAY button. -Select your PhenolRedLactose medium sample tube. -Click and Record Results. Observations and Results Gram stain result: Experiment 1: Pink rods and the first stain showed negative rods. Experiment 2: Case Study 67 Rods were gram positive cocci. Blue circular stains. Case Study 161 were negative pink rods. Part I: Contamination • Step 6: What color are the traffic lights after caps are removed? Contamination is yellow. • Step 7: What color are the traffic lights after caps have been left off and then replaced? Once left off and replaced the inoculated light is green and the contamination is Red. • Step 9: Appearance of tube: Red • Step 12: Appearance of tube after overnight incubation: Cloudy and Yellow. Part II: Aseptic Transfer Tube-to-Tube Test • Was your aseptic transfer successful? (contamination light and Inoculation light both green) Yes • Appearance of tube after overnight incubation: Remained yellow and cloudy Discussion -What precautions are taken in a "wetlab" to prevent contamination? Label all items Wear proper clothing (gloves/lab coat) Sterilize all equipment and materials Clean up any and all spills properly. • During the Contamination portion of the exercise, why was the contamination not present when you first placed the phenol red glucose tube in the 37° incubator? Incubation may not have been carried out the proper amount of time to see the phenol red glucose reaction. • After your sample had been incubated for 1 day, why is the solution turbid? What did the color change in the medium indicate? The incubation period was successful. The change in medium indicates the growth of the cultured organisms. • What caused the contamination of your sample? • How successful were you with the aseptic technique? How many times did you need to repeat the experiment until you were able to successfully inoculate your media without contamination? You must follow all policies and procedures to keep from contaminating your area and/or your sample. You must maintain sterilization of your instruments, all inoculation tools and always wear eye wear for your safety. Solutions must remain in the autoclave for sterilization. All pure cultures are opened in sterilized areas. Repeated trials the same way should give same and accurate results. If all policies and procedures were not followed, contamination could occur. • How important is sterile technique in a wetlab? Why? If you don’t have a sterile technique in your wetlab it’s possible to give false positives, which could skew results and affect things negatively. Could also cause unwanted growth/cultures on the mediums which would also lead to contamination. Conclusion In conclusion I was able to determine if there was any contamination or spread of pathogens by following the procedures given and doing the experiment as directed. By following all procedures I was able to keep my wetlab sterile and by my gram stains showing negative rods, I was able to confirm that there was not any contamination. I was able to determine this based on the colors of the medium present. BACTERIAL ANATOMY Introduction Include purpose of lab experiment Brief summary of topic investigating and case study State major finding Procedure -Get an Unknown -Perform Biochemical Tests -Identify Bacterium -Review Results Experiment 2: Bacterial Anatomy -Type Anatomy 1 in the “Enter a Label” line. -From the subgroup dropdown menu, select Bacterial Anatomy 1. -Click Auto-Inoculation allowed. Click OK. -Record details of your case study scenario. -Observe the morphology of the bacteria and record results of gram stain. Select Bacterial Anatomy 2 Exercise: -Click the NEW UNKNOWN and label the subgroup. -Follow the same instructions for the first Bacterial Anatomy experiment. Observations and Results Gram stain result: List Gram reaction and shape (e.g., Gram-positive rod) (Experiments 1 and 2) Discussion In the Discussion section, answer the following questions. • Describe the morphology of each bacterium (e.g., cocci, rods, or spiral or curved). Coccus: Spherical -Oval, Elongated or Flattened Bacillus: Rod-Shaped -Single rods//after division they can be in pairs Spiral: Twisted -one or more twists (The size, shape, and arrangement of bacterial cells. Retrieved November 2, 2016, from • Describe the arrangement of the bacteria. Were the bacteria organized as singles, pairs, chains, clusters, or another type of arrangement? Bacillus can be in pairs only after division. Spiral can be in clusters, in groups of one or more twists. • Describe the Gram stain procedure and how to interpret the results. How does this apply to your Sample 1 and Sample 2? Which stain is responsible for the results that you see? • Relate the results to cell structure. You can also use Internet resources. Make sure that you cite all external data sources, including your textbook. • Indicate the terminology that is used for the cell anatomy and arrangement observed for the organism in Sample 1 and Sample 2. Conclusion In a few sentences, provide a concluding statement about the results of your lab.