BIOS 242 Week 2 iLab: Aseptic Technique
BIOS 242 Week 2 iLab:
BIOS 242 Week 2
BIOS 242
BIOS 242 Week 2 iLab: Aseptic Technique
ASEPTIC
... [Show More] TECHNIQUE
Introduction
The purpose of this lab is to culture cells successfully without contamination using aseptic technique. If contamination occurs, the lab experiment will be inaccurate. Biological contamination can happen via dirty supplies, airborne toxins, and unkempt workstations. This aseptic technique lab is designed to guide you through a successful (uncontaminated) lab experiment. With these aseptic techniques it will help to yield a level of protection for the cell culture. Aseptic techniques require sterile media, sterile agents, proper hygiene, and a clean workspace.
Findings: Mannitol Fermentation test 100% positive.
Procedure
Conduct Contamination Test:
1. Click the New Unknown button. A window will open asking you to enter a
label and select a subgroup.
2. Type Technique1 in the “Enter a Label” line.
3. From the Subgroup dropdown menu, select Asceptic Technique. DO NOT
CLICK Auto-inoculation allowed. Click OK.
4. Record details of your case study scenario.
5. Record the Gram Reaction by clicking the appropriate radio button. Click
Record Gram Reaction.
6. You will need to indicate the results of the Gram Stain in your Lab Report.
7. From the Media dropdown menu, select Phenol Red Glucose Broth Durham
Tube. The Media dropdown menu is to the far right of the New Unknown
button.
8. Enter PhenolRedGlucose in the Medium Label window to label your sample.
Click OK. Two tubes will appear on your workbench. Take a careful look at
the sample tube (the one on the right.) Is there media in the Durham Tube
(this is the tube that is upside down in the medium.)
9. Note the status of the Traffic Lights for Inoculation and Contamination in the
upper right hand corner of the lab software window.
10. Right click on the phenol red tube (the tube on the right) and select Remove
caps/lids.
iLab: Aseptic Technique and Bacterial Anatomy and Morphology Page 4
11. Look at the Inoculation/Contamination traffic lights. What color
are these lights?
12. Right click on the phenol red tube and select Replace caps/lids. Look
at the Inoculation/Contamination traffic lights. How has the color
of each light changed? Record these changes for your lab report.
13. Using your cursor, drag the phenol red tube and place it on the 37
degree incubator (this is the incubator on the right side of the
workspace. Notice the reading on the thermostat). This places the tubes
in the incubator. You will find that the tubes disappear from the desktop.
14. Right click on the 37 degree incubator and select PhenolRedGlucose.
The tube is now on your workspace.
15. Observe tube.
16. Replace tube in 37 degree incubator. Note that the Virtual Days counter
is 1.
17. Click New Day. The Virtual Days counter now reads 2.
18. Retrieve tube from incubator. How has the appearance of the contents
of the tube changed? Is it still clear? Is it cloudy (turbid)? If the
sample is turbid, then this means that bacteria has grown in the media.
What is the source of this bacteria?
19. Right click on tube and select Record Results. For this experiment,
record a POSTIVE result for both Acid from Glucose and Gas from
Glucose tests. Note that when you record results that your sample is
no longer on your workspace. This is because it has been automatically
been transferred to the biohazard bag and discarded.
Observations and Results
Gram stain result:
Gram negative and Rod shaped
Gram reaction {-} Eliminated {53} Remaining {71} Recorded 2 times
Acid from glucose {+} Eliminated {0} Remaining {71} Recorded 1 time
Gas from glucose {+} Eliminated {15} Remaining {56} Recorded 1 time
Lactose fermentation {+} Eliminated {25} Remaining {31} Recorded 1 time
Part I: Contamination
• The color of the traffic lights after caps are removed were yellow.
• The traffic lights turned red after caps have been left off and then replaced
• The color of the sample has now change to yellow and it is turbid. The source of the bacteria is staphylococcus aureus.
Part II: Aseptic Transfer Tube-to-Tube Test
• Was your aseptic transfer successful: Not on the first attempt
• Appearance of tube after overnight incubation: The culture changed from yellow to red
Discussion
Wetlab precautions:
Cleanliness is the number one precaution while preforming this experiment. In order to ensure no contamination occurs, you must first set up hood/area properly. Also make sure there is experiment is not near drafts because that will blow dirt into your area. Clear all clutter from workspace. Make sure you used 70% ethanol as your cleansing agent. Make sure to sterilize tools before and after experiment.
In terms of the experiment there was no contamination present when I first placed the phenol red glucose in the tube because the contents had the reach the correct settings and temperature for the contamination to oocur; also timing played a big role in contamination too. So, after a day the solution became turbid because it had time to ferment and contaminate. The color change of the medium indicate that it was contaminated. The contents had mannitol fermentation and tested positive for staphylococcus aureus. I had to repeat the experiment several times before I was success at inoculation of the media without contamination. If contamination occurs, the wet lab experiment will be inaccurate providing false results.
Conclusion
Aseptic techniques require sterile media, sterile agents, proper hygiene, and a clean workspace Biological contamination can happen via dirty supplies, airborne toxins, and unkempt workstations. The aseptic techniques lab are designed to guide you through a successful (uncontaminated) lab experiment. With these aseptic techniques it will help to yield a level of protection for the cell culture.
BACTERIAL ANATOMY
Introduction
The purpose of the bacterial anatomy experiment is to understand the anatomy of bacterial cells. This experiment is designed to find different structures that are linked to bacteria.
Case study involves: A gal named Denice. Denice does an experiment centered around how the skin microflora changes in immunocompromised hamsters. She used both compromised and normal hamsters as her test subjects in the same environment.
After two weeks both hamsters had the same organism found on their skin.
Procedure
Steps taken for this lab:
• Type Anatomy1 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 the results of the Gram Stain.
Observations and Results
Experiment 1.
Gram stain result: Gram positive cocci. Gram reaction- {+} eliminated {71} Remaining {53} Recorded 2 times. Gram shape-clusters and dark purple and light purple in appearance.
Experiment 2.
Gram stain result: Gram negative rods. Gram reaction- {-} eliminated {71} Remaining {53} Recorded 1 time. Gram shape-scattered elongated rod shaped. Orange in color.
Discussion
In experiment one of the bacterial anatomy the morphology of the bacterium resembled small circular balls and in clusters. The color of the staining were purple and light purple. In experiment two the bacterium resembled rod shapes and orange in color; some of them were clustered together while others were single.
Gram stain procedures are:
First label the slide, prepare specimen, heat the slide, put slide on rack, use with crystal violent solution, rinse slide with water, place gram iodine on slide, wait 60 seconds, rinse slide with water, drench slide in decolorizer until no violent color is present, no wash the slide off with water, now put safranin on the slide, rinse with water, and finally gently dry the slide with bilbous paper towels until it is dry. A thick and single cell wall will mean that it is gram’s positive bacteria stain and a thin multiple layered cell wall will mean that it is a gram’s negative bacteria stain. Sample one is a perfect example of gram’s positive bacteria and sample two is an example of negative bacteria stain.
Conclusion
Understanding the bacterial anatomy is helpful because it can be used as a method to preventing mass sicknesses and deaths. [Show Less]