Wednesday, October 22, 2008

The more legs you have, healthier cells you'll get
The shining dendrites show that the cell are dividing

The cell has changed into multipolar morphology

The confluent melanocytes after day-12

This is the story about my cell. I used to work with melanocytes or ‘mel’ the other name given by me. At first glance, I don’t have any idea on how difficult to handle melanocyte. In early of 2007, I bought the first vial of normal human melanocytes from Cascade Biologics. I can’t describe about what I feel for the first time to handle this cell. Before play around with melanocyte, I used to handle 3T3-L1 supplied by our senior Ting. For the first timer, I quite easy to handle melanocyte based on some knowledge from journals and articles. But I did a big mistake on the first step to thaw the cells. I used to follow the general procedure that being adapted on cell thawing. I forgot, that my primary cell is different from other cell line that we have in the lab. We can’t centrifuge normal cells. We just add medium then let the cells grow about 72 hours without disturb. For my first batch, the first and second passages are growing as expected. The cells grow beautifully with dendrites attached to the flask. I observed the cells, and then I know the cells grow from the dendrites. For melanocytes, the more legs you have, the healthier cells you get. I also get to know, the healthy cells are shining like a star. Suddenly, I start to have problem with the cells. I have a lot of dots in the flask that I can’t figure out whether it is contaminated or it just the byproduct from the cells. But the medium get cloudy and that moment I don’t have any experience to handle problematic cells. I could not safe my cells and even to finish my standard growth curve. Then, with the generosity of CEPP, I bought the second vial of melanocyte. At this time, the scenario is different. I was given a dead cells vial to thaw. Luckily, I have got my replacement from the principal. For the third vial, my heart still pumping fast like I never do the thawing procedure before because my cell is very sensitive. Form the findings, normal human melanocytes has passage limitation. Adult melanocytes can grow until 5 to 6 passages meanwhile foreskin melanocytes can survive until 7 to 8 passages. For the third vial, I managed to analyze the melanocytes for melanin and tyrosinase assay. I used to handle the cells on my own routine based on my experience and observation. When I see strangers in my flasks, I will increase the percentage of penicillin/streptomycin up to 2%. I need to change medium every 24 hours. The melanocyte is very sensitive, so PBS is restricted to wash the medium or else you will loose the cells. I used to wash using the medium so it makes the maintenance costly. You always feel difference when working with melanocyte. The best part handling melanocytes, when I have to talk with them and cuddle like my own precious baby. It works, even though they can’t survive till the end. When handling melanocytes, you have to shut off the light because it is sensitive to light. Don’t be shocked, the pellet of melanocyte is black in colour meanwhile most of the other cell pellet are yellowish. The cell count also not so exciting, the most I can reach is almost the power of 6. Many studies have been done using melanocytes, but most of researchers are prefer to use melanoma instead of using melanocytes because of melanocytes limitation. Whatever it is, it is wonderful to work with melanocytes and it teaches me how to be patient and think fast how to overcome the strange situation.

1 comment:

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