For slants, we recommend using screw capped tubes. For cultures on Petri dishes, the plates need to be sealed with Parafilm. Sealing the plates not only helps to prevent molds from sneaking into the plates, but it slows the agar from drying.

Optimal Growth Conditions Thus, a microbiologist will incubate a particular strain of bacteria at its optimal temperature so that he can study it when it is healthy. Organisms that grow best at human body temperature, which is approximately 37 degrees Celsius (98.6 degrees Fahrenheit), are called mesophiles.

Two incubation temperatures and times are used: 37 °C for 24 hours to encourage the growth of bacteria of mammalian origin, and 22 °C for 72 hours to enumerate bacteria that are derived principally from environmental sources.

At 35°C, most human bacterial pathogens with differing optimum growth temperature will grow reliably, although colonies may appear small or require additional incubation due to their slower growth rate. Incubation at 37°C also risks fluctuations to dying off temperatures.

Some bacteria cannot be grown with nutrient agar medium. Fastidious organisms (picky bacteria) may need a very specific food source not provided in nutrient agar. One example of a fastidious organism is Treponema pallidum, bacteria that causes syphilis.

Checking for Contamination Look for signs of fungal contamination. Fungal contamination will appear as fuzzy, filamentous, or hair-like growths, and should be visible to the unaided eye. Fungal contamination often occurs right along the edge of an agar plate. Inspect for signs of bacterial contamination.

make a desired broth solution and cut a small portion of slant containing the organims. let it grow for 24-18 hrs and there after sonicate it for 30mins only in a sonar bath not in the probes, and incubate it at 37 C for 24-48 hrs. Then follow streaking method on a nutrient plate and repeat it for several times.

The subsequent recovery of cryopreserved cells is straightforward: Cells are thawed rapidly in a water bath at 37°C, removed from the freeze-medium by gentle centrifugation and/or diluted with growth medium, and seeded in a culture vessel in complete growth medium.

Cover the ampoule with a sterile cotton sheet, and cut it carefully at the neck. Do not use a cotton sheet containing alcohol. Using a sterile Pasteur pipette, add 0.3 to 0.5 ml of suitable rehydration fluid into the ampoule. Spread the sample on a suitable plate and incubate it under the directed condition.

reviving the strain Add aseptically approximately 1ml nutrient broth with at sterile Pasteur pipette to the freeze-dried material and mix well. Leave the material to rehydrate for five-fifteen minutes. Transfer a few drops of the suspension to an agar plate and spread (purity check).

To recover bacteria from your glycerol stock, open the tube and use a sterile loop, toothpick or pipette tip to scrape some of the frozen bacteria off of the top. Do not let the glycerol stock unthaw! Streak the bacteria onto an LB agar plate. Grow your bacteria overnight at the appropriate temperature.

Thaw frozen cells rapidly (< 1 minute) in a 37°C water bath. Dilute the thawed cells slowly before you incubate them, using pre-warmed growth medium. Plate thawed cells at high density to optimize recovery. Always use proper aseptic technique and work in a laminar flow hood.

Thaw the tube containing the frozen cells in a 37°C water bath for 2 minutes. Immediately transfer the thawed cell stock to the flask containing the equilibrated growth media. Incubate cells overnight at 37°C, 5% CO2 and replace media the next day. Continue to incubate the revived cells for 48 hours and change media.

It is vital to thaw cells correctly in order to maintain the viability of the culture and enable the culture to recover more quickly. Therefore, it is essential that cultures are thawed quickly and diluted in culture medium to minimize the toxic effects.

For the 2nd question, fast thawing is needed to avoid recrystallization during rewarming process (small crystals can grow bigger and cause ice injury to cells during rewarming process). Therefore, we simply heat the samples through this region as fast as possible.

Freezing usually damages cells because water expands when it freezes. Animal cells just have thin membranes around them. When ice crystals form, they destroy the cells. That’s what frostbite is.

Freezing does not kill germs and bacteria. Instead, it essentially puts them into hibernation. They are inactive while the food is frozen and will “wake up” as soon as the food thaws. And as the food thaws, so will the moisture, which means the bacteria will have the moisture it needs to survive.

Advantages and disadvantages of cell culture The main advantage is the consistency and reproducibility of results that can be obtained from using a batch of clonal cells.

Henrietta Lacks and her “immortal” cells have been a fixture in the medical research community for decades: They helped develop the polio vaccine in the 1950s; they traveled to space to see how cells react in zero gravity; they even aided in producing a vaccine and reducing HPV infections—and subsequently instances of …

HABERLANDT

Disadvantages of Animal Cell Culture Ø Chances of chemical and microbial contamination are very high in in vitro methods. Ø High possibility of cross contamination of different types of cells in culture. Ø Experience and expertise are required for an effective maintenance most of the cells.

Among the weaknesses of cellular models is the inability to reflect the role of complex decision making by human agents. However, cellular frameworks may be combined with Markov models, in which transition rules are treated probabilistically and may be conditioned by temporal lags in cell response.

2.2 Cell-wall PMWs Wood-destroying organisms do not degrade them and they are highly resistant to acids and bases. However, cell-wall PMWs are more brittle than the parent wood cell wall, which is their chief disadvantage.

Mitochondria are the part of the cell that produces ATP which the cell can then use as energy to do work. Muscle cells require a lot of ATP to carry out contraction and therefore a large number of mitochondria are required to produce this ATP.