User:Madeleine Y. Bee/Notebook/CHEM-572 2014S/2014/04/22

Cell Tests

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April 22, 2014 Objectives Cell tests (only 2&3) Test Procedures Type I Collagen Test (Detection and Quantification) Company procedure Collagen Staining Procedure & Microscope Imaging Company procedure Trypsin Cell Detachment Procedure & Cell Counting Company procedure Test Results Collagen Staining Procedure & Microscope Imaging Test Absorbance data: Calculations (from company procedure) "To calculate the amount of collagen, correct the OD 540 value by subtracting the contribution of Fast Freen at 540nm, which is 29.1% of the OD 605 value. The color equivalence (OD values/ug protein) is 0.0378 for collagen and 0.00204 for non-collagenous protein at OD 540 and 605, respectively." Collagen (ug/3 scaffold layers)= [OD 540 value-(OD 605 value x 0.291)]/0.0378 Non-collagenous proteins (ug/3 scaffold layers)= OD 605 value/0.00204 Trypsin Cell Detachment Procedure & Cell Counting Performed on 3 scaffold layers. “Original” - 125 cells on scaffold Polydopmaine & “original” - 450 cells on scaffold Polydopmaine only- 150 cells on scaffold Brute force - 300 cells on scaffold Plain - 100 cells on scaffold Next time Perform all three tests in triplicate Have one scaffold for assays that has no cells one it (for blank & with "leftover" media [under microscope?]) Keep solutions post-assay to look at under microscope to see if cells are still there