Chang Lab:Notebook/CBE/08/149/2008/12/18

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 * style="background-color: #FFF"|   Antimicrobial efficacy testing of antibiotic-containing biodegradable nanopolymers against biofilm and planktonic cells
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 * style="background-color: #F2F2F2" align="center"|  |Main Notebook page


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Logs

 * Start 9:15 AM
 * Took 5 ml from 10-ml overnight E coli culture into 150 ml broth for 5-6 hr incubation at 37 C (inoculum for biofilm optimization trial III)

Biofilm Optimization (Trial III)

 * 3 CBDs are prepared to be incubated at 3 different temperatures: 21, 30 and 37 C at 30 rpm for 24 hours. The procedures below follow from previous trials, except minor adjustments to procedures in: M9 inoculum resuspension sequence => broth matched with 1.0 McFarland first before centrifuging.
 * Following 1.0 McFarland Std (A600 = 0.257), 5-hr incubated E coli broth adjusted to match A600 value. Approximately 9X (0.3ml broth culture to 2.7ml broth) dilution to match standard.
 * For minimal medium M9, absorbance of E coli broth at 600nm adjusted until it matched 1.0 McFarland Std. E coli 1.0 McFarland inoculum centrifuged at 3000g for 10 minutes, supernatant LB Broth discarded through careful pipetting, without disturbing sedimented E coli cells. Previously prepared M9 medium used to resuspend E coli cells, same volume at 1ml.
 * Using multipipette (8-pronged), 200 uL of adjusted inoculum pipetted into each well of CBD.
 * Columns 1-6: rich M9 Medium
 * Columns 7-12: minimal LB Medium
 * CBD covered with pegged lid. Incubated at desired temperatures at 30 rpm. Will check on 12/19 to score for biofilm growth.

Serial Plating

 * Plated LB-based 1.0 McFarland E coli inoculum at 10^-10 to 10^-20 dilutions to verify cell number.
 * Plated M9-based 1.0 McFarland E coli inoculum at 10^-6 to 10^-10 dilutions to verify cell number.
 * Plated planktonic cells from CBD wells at 10^-2, 10^-4, 10^-6 dilutions for both LB and M9 wells of 21, 30 and 37 C CBDs.
 * To be checked after ~24 hrs for colony counting.


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