04 May, 2009

Bloggable Science-Like Paste

I did a little quasi-scientific quasi-experiment for your entertainment. I'm curious to see how quickly all of you can poke holes in it.

Informal Background:
So, the other day I found a sleeve of plates in the fridge that were rather old and on the verge of becoming trash. My boss has told us that if we need to keep our work sterile but can't fit it in the flow hood (or someone else is in the flow hood already), we can just light a Bunsen burner and it'll create an updraft that sweeps all the microbes away from our work. I don't trust my cell culture media with this, but I am willing to use this for bacteria work. However, I haven't seen this qualitatively proven with my own eyes or in the literature so I figured I should test this.

Hypothesis:
A burning Bunsen burner is sufficient to keep your work sterile within 1m.

Materials and methods:
Set up line of split TSAII/5%SB and MacConkey agar plates from BD. Lit Bunsen burner and allowed to burn freely for 5min. Then turned over plates and removed lids from closest to farthest. Plates were then left uncovered for 5min and re-covered while the Bunsen burner burned. Plates were incubated 48h at 37C, high humidity, 10% CO2 in a water-jacketed incubator. After 48h, bacterial growth was assessed.

Figure A: Toaster's benches showing experimental setup. My benches are particularly messy here because I was also running an ELISA at the same time and did this during an incubation period.

Results:
8 out of 10 plates remained sterile. The 2 plates that did not remain sterile were plates 1 and 3 (numbered going out from Bunsen burner). Growth on both of these plates was a single large colony of yellowishness on the blood agar side.

Conclusion:
Inconclusive. Growth on plates did not follow distance as one would logicaly expect if the Bunsen burner does indeed create an updraft. Growth that was observed was likely Gram positive (because it didn't grow on MacConkey agar, which contains bile salts that inhibit Gram positive organisms' growth).

2 comments:

tig said...

Could be anything, bile salts don't just inhibit Gram positives. They inhibit lots of stuff. Also, many Gram negative bugs can grow on blood agar but won't touch MacConkey's.

I use a Bunsen for all aseptic work. Never used a flow hood and never intend to. I've subcultured Bacteria to-from brain-heart infusion agar and broths (BHI is, in my opinion, the easiest medium to contaminate cuz everything just seems to love it) using just a Bunsen and I think I've had many 2 contamination issues in the last 5 years.

Ambivalent Academic said...

There is likely to be an outer limit to the updraft...in which there is a complementary down draft. Methinks your plates 1 and 3 were in the downdraft zone, as most people who rely on the updraft principle for sterility actually open their plates/vials/whathaveyou rightnextto the flame. Even leaving plates on the bench directly under the burner is too far away to take advantage of the updraft. Or so says conventional wisdom around these parts anyway.