Do not set the cap down and ensure that the open end is facing downward. Do not lay the loop down! Now you may lay the loop down until it is needed again. Transferring the inoculum to an agar plate: lift the edge of the lid just enough to insert the loop. Test for coolness by touching the agar at the edge of the plate. Pick up a loopful of liquid inoculum or bacterial growth from the surface of an agar plate and, starting about 25 mm in from the edge of the plate 1 , streak lightly back and forth with the loop flat, making close, parallel streaks back to the edge of the plate.
Flame and cool the loop again, and make another set of streaks starting from 3 perpendicular to and crossing the second set of streaks, but avoiding the first set. To prevent the transfer of saliva. By using solid thermal insulators.
Found in Genesis , mighty cherubim and a flaming sword were placed at the entryway to prevent their return. Lets say theirs two cultures, culture A and culture B, the people of culture b believe that culture a do some action that is voodoo in culture b. The easiest way to prevent biomagnification would be to cut down or better yet eliminate our pollutants so they cant transfer. You want to keep the talking down to a minimum during the first few days just to prevent en flaming your tongue further.
It is to prevent contamination of the culture from airborne microbes. To kill excess organisms to reduce the chances of contamination and to prevent mixing different bacterial colonies.
This actually sterilizes the mouth of the test tube. It is a back up to prevent contamination. Log in. C Programming. Add an answer. Want this question answered? Study guides. Q: How can you prevent splattering when flaming the loop which has just been used to transfer a culture? Write your answer Related questions. You will be working with many pathogenic species of bacteria in the laboratory. Remember that bacteria are in the air as well as on the skin, the counter, and all objects and equipment that have not been sterilized.
The most important tool for transferring cultures is the wire inoculating needle or loop. It can be quickly sterilized by heating it to red hot in a Bunsen burner flame. Adjust the air inlets of the burner so that there are a hotter inner cone and the cooler outer flame. A dry needle may be sterilized by holding it at a degree angle in the outer part of the flame.
A wet loop with bacteria on it should first be held in the inner part of the flame to avoid spattering, and then heated until red hot in the outer part of the flame. Always flame the loop immediately before and after use! Allow it to cool before picking up an inoculum of bacteria or you will kill the bacteria. Hold the loop or wire handle like a pencil. Mixed cultures more than one species can be isolated using the streak plate technique.
The goal is to acquire a pure culture of one species of bacteria, in a single colony, from a mixed culture We do this by separating the microbes on the surface of agar with quadrant streaking, this method DILUTES the bacteria. Do not allow pipettes to come into contact with corrosive chemicals. Although working within the sterile field created by the Bunsen burner, do not flame micropipettors, tubes and plastic tips.
The tubes and tips should be pre-sterilized. The micropipettors may be wiped down with a pre-moistened disinfectant wipe prior to use. A numeric volumeter showing the dispensed volume can be set by turning adjustment knob. Adjust the volume before proceeding to step 3. To obtain maximum accuracy when decreasing the volume setting on the micropipettor, slowly dial down the thumb wheel, making sure not to overshoot the graduation mark.
Then slowly dial down the thumb wheel to reach the intended volume, making sure not to overshoot the graduation mark. The volumeter shows three numbers. Depending on the micropipettor, the numbers are interpreted differently. Note that each micropipettor is only as accurate as the smallest graduation mark.
P2: For volumes between 0. The top number denotes volume in microliters. The second number indicates tenths of a microliter 0. Each graduation mark equals an increment of two one-thousandths 0. P For volumes between 1. The top number is for tens of microliters; this usually is set at "0" and should only be set at "1" with the other two numbers set at "0" when dispensing The middle number denotes volume in microliters. The third number indicates tenths of a microliter 0. Each graduation mark equals an increment of two one-hundredths 0.
P For volumes between 2. The top number in black is for tens of microliters; this should only be set at "2" with the other two numbers set at "0" when dispensing The second number in black denotes the volume in microliters. The third number in red indicates tenths of a microliter 0.
P For volumes between The middle number indicates the dispensed volume in tens of microliters, and the third number denotes volume in microliters. Each graduation mark equals an increment of two one-tenths 0. The middle number is for hundreds of microliters.
The bottom number is for tens of microliters. Use an analytical scale to measure water, making sure the minimum and maximum settings correspond to the intended volume. Since water has a density of 1, then 1 ml of water is equivalent to 1 gram g.
Also, make sure the tip does not leak and can maintain the desired volume until dispensed using plunger system. Micropipettors must be used with plastic disposable tips at all times.
Fit a tip tightly onto the end of the barrel of the micropipettor. Press down and twist slightly to ensure an airtight seal. Tips are usually packed into plastic boxes that can be sterilized by autoclaving.
Open the tip box to retrieve a tip, then close the tip box to minimize contact with contaminants in the air. Some tips have filters similar to the cotton wool plugs on serological pipettes. These tips are often more expensive than regular tips and thus are used for specialized applications.
For instance, when measuring volatile chemicals such as chloroform or radioactive liquids such as 32 P-labeled DNA, using filter tips helps prevent the barrel of the micropipettor from getting contaminated. Keeping the micropipettor upright will prevent liquids from running inside and contaminating the barrel of the micropipettor. The micropipettor has three positions: 1 Rest position, 2 First stop, and 3 Second stop Figure 6 , panel B.
The instrument has a two-stop plunger system. The first stop has two functions. The first is to draw in the desired volume of liquid into the tip when releasing the plunger from the first stop to the rest position.
The second function is to dispense the majority of liquid from the tip when depressing the plunger from the rest position to the first stop. Further depressing the plunger to the second stop dispenses whatever liquid remains in the tip. Depress the pushbutton on the plunger from the rest position to the first stop. Air equal to the volume of the setting will be displaced.
Do not touch the micropipettor itself to the sides of bottles, tubes and flasks; otherwise the inside surfaces of these vessels will become contaminated. Only the tips are sterile. Release the pushbutton slowly to aspirate the liquid into the tip. Stop once the pushbutton is back to the rest position. Wait a moment so liquid can be drawn into the tip. The volume of liquid in the tip will equal the volume of the setting of the micropipettor.
Remove the tip from the liquid, and visually inspect the tip to confirm that the liquid drawn up has reached the expected level in the tip and there are no air bubbles in the tip. If necessary, expel the liquid and manually tighten the tips onto the micropipettor. Draw up the liquid and check again. To expel the liquid, slowly depress the pushbutton on the plunger to the first stop. Wait a moment then press the pushbutton to the second stop to expel any residual liquid in the tip.
Depressing the plunger too quickly may cause the liquid being expelled to splatter or will produce undesirable bubbles in the tube. Discard tips into designated sharps waste container by pressing the ejection button on the micropipettor.
When finished with an experiment requiring use of aseptic technique, turn off the Bunsen burner, then put away all supplies and reagents. Wipe down the outside surfaces of labware bottles, micropipettors, pipette tip boxes with a pre-moistened disinfectant wipe to ensure contaminants are not transferred to the storage location.
Place contaminated glassware and hazardous waste materials into the proper disposal receptacle. Laboratory waste includes labware such as gloves, pipettes, tips, and tubes.
Non-infectious hazardous waste is generated when performing experiments with non-pathogenic organisms BSL-1 while infectious hazardous waste is generated when using pathogenic organisms BSL-2 or above. Infectious waste must be autoclaved or disinfected before it is discarded. Wipe down the entire work area on the laboratory bench with a pre-moistened disinfectant wipe from the canister, once again allowing the disinfectant to evaporate.
Wash hands thoroughly with antiseptic soap and warm water before leaving the laboratory. A sample application for using serological pipettes to transfer liquids is shown in Figure 7. These pipettes often are used in the microbiology laboratory to prepare media for inoculation with bacterial cultures.
For example, sterile flasks first are filled with a specified volume of culture broth, in this case Luria Broth LB , then a small number of cells such as E. Using a serological pipette, first the broth must be aseptically transferred from the media bottle to the flask.
In this case, 25 ml of LB was added to a ml sterile flask using a 25 ml serological pipette. Next, the broth must be inoculated with E.
The flask is incubated in a growth chamber for a particular amount of time, allowing the cells to replicate for this example, the E. The result is a turbid bacterial cell culture that can be used for subsequent experiments.
Serological pipettes also may be used to transfer media originally supplied in a bottle to test tubes, or between test tubes, as is done when making dilutions of a bacterial culture. If aseptic technique is not maintained throughout these types of media manipulations, then cultures will become contaminated, and subsequent experiments using those cultures will be delayed because fresh, uncontaminated cultures will need to be prepared.
Errors occur because a sterile field is not maintained throughout the procedure. For instance, you may forget to disinfect the laboratory bench or flame the rim of a culture bottle or tube.
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