Sunday, November 18, 2007

Results of my MTT assay.

Hey guys ive been talking alot on MCT stuff and MTT assay. Now its time for me to show the fruits of my labour...........the RESULTS!!!

Cell Sensitivity (MTT) assay

Cool ah?? Hahha. With this result, i will find the IC50 of the drug against the Hep G2 cells.

IC50 or the half maximal inhibitory concentration, represents the concentration of the inhibitor (drug) that is required for 50% inhibition of its target (Hep G2 cells). IC50 measures how much the drug is required for 50% inhibition in-vitro.

I will take a minimum of 5 points to form a linear line. Then using Microsoft Excel, i will obtain the equation of line and calculate the IC50.

With the equation, substitute the y value with 50 and then the x value could be obtained and that is the IC 50 of drug towards Hep G2 cells.

Enlightened?? Hahhaha See you guys in sch.

Najib Bin Hamid

Monday, November 12, 2007

Preparation of a Hanging Drop

this is a simple experiment to observe the movement of microorganisms under microscope.

- bacteria sample
-cover slip
-glass slides
-inoculating loop

  1. Use plasticine provided; make a loop about the size of a 5 cent coin.
  2. Place the loop firmly onto the center of a clear glass slide.
  3. Aseptically transfer a loop of sample broth culture onto the center of the clean cover slip.
  4. Place the glass slide (with the plasticine loop facing downwards) over the cover slip and turn the glass slide over. The drop of culture must appear to hang down from cover slip.
  5. View 10X and 40X
Movement of E. Coli was observed. i thought the E. Coli will be swimming, but actually movement of E. Coli under microscope is just vibrating in the original position.

Difficulty faced during experiment is when i was expected to turn the glass slide over, the E. Coli tends to slide to the side of the plasticine. it was only after a few tried that i finally managed to do it right.

that's all!

Suat Fang

Friday, November 09, 2007

Plant subcultuing

Hi guys! I'll be blogging on something that we do not usually have the chance to do in our curriculum! During this period, I'm like a gardener, the only difference is that I'm working in a plant lab and not a garden.. :)

Okie! Now about plant subculturing...

A subculture is actually a culture made by transferring tissue from a previous cultures to a fresh medium in order to prolong the life of a particular strain where there is a tendency to degeneration in older cultures. A plant subculture in many ways are similar to a cell subculture in terms of aseptic techniques involved, characteristic of a subculture and precautions to be taken.

Before a subculture can be carried out, the media used should be autoclaved at 121 deg cel for 15 min in order to kill off any microorganisms. After the media is autoclaved, the container should not be open unneccessary as it is prone to contamination. If necessary, it can only be opened in a horizontal flowhood which confers more product protection.

The selected plant culture has to be in a healthy state and free of contamination. The whole subculturing process have to be done in the horizontal flow hood to minimize contamination. Forceps and scissors should all be sterilized before use. The flow hood should also be UV and swapped with 70 % ethanol before use. Strict aseptic techniques applies. The containers is opened by applying force onto the four edges of the containers with thumbs and index fingers only. Once the plant container is opened, hands with gloves should not be placed over the plant culture. Only sterilized scissors and forceps are allowed to touch any of the plant culture, media or within the containers. The lower portion of the stem are cut and retrieved out using a forcep. All the leaves are then removed, using 2 forceps. Hence by removing the leaves, the nod of the stem is exposed in which new plantlets will grow from them. The stem is then inserted lying horizontally into the media with the use of forceps. the stems are aranged in organized manner (in rows) so as to prevent overcrowding.

Finally the plant subculture is ready for growth in the growth room. Nevertheless, the plant subculture are supplied with 12 hours of light daily and should be constantly check for contamination. Once sufficient plant subculture is grown, it is havested for use.

feel free to ask any question!

Royston Tan