Sample 1 + Lysozyme extraction mixture (with dH2O) of egg + bacteria These Abs i
ID: 151521 • Letter: S
Question
Sample 1 + Lysozyme extraction mixture (with dH2O) of egg + bacteria
These Abs is going down. (delta Abs > 0)
But, after this first extraction mixture,
All of Abs is going up. (following sample 2~8) (delta Abs < 0)
Sample 2 + Lysozyme soultion with pH8.2 Tris EDTA buffer by (Ultra)-filtrated from sample 1 + bacteria Sample 2 + Lysozyme soultion with pH8.2 Tris EDTA buffer by (Ultra)-filtrated from sample 1 + bacteria
Sample 3, 4, 5 + fractions of sample 2 by ion-exchanger with pH 8.2 Tris EDTA buffer + bacteria Sample 3, 4, 5 + fractions of sample 2 by ion-exchanger with pH 8.2 Tris EDTA buffer + bacteria
Sample 6, 7, 8 + fractions of sample 2 by ion-exchanger with pH 10. 5 Carbonate buffer + bacteria Sample 6, 7, 8 + fractions of sample 2 by ion-exchanger with pH 10. 5 Carbonate buffer + bacteria
What is wrong with this Enzyme assay? And I got the top level of the purification from sample 6. (Lysozyme pI.11) Sample Table-[Active ] Ex Conc WL450.0 Sample IlD 30 60 90 120 150 180 210 Type Unknown Unknowrn Unknown Unknown Unknown Unknown Unknown 0.77243 0.80312 0.83733 ettt0.86934 0.89392 0.90878 0.91829 2 4 6 7
Explanation / Answer
Lysosome assay is used to test the ability of the enzyme to cleave bacterial cell wall. The lysoyme activity is enhanced in the presence of EDTA tris buffer. This EDTA chelates the metal ions needed for cell's survival. Thus the cell is unable to obtain these metal ions and lysosome cleavage will be easier. Thus , as the EDTA -lysosyme concentration increases, the turbidity of the solution incraeses . Due to increase in turbidity the absorbance values decreases. But here the absorbance bvalues increases. This shows that teh cells are not lysed by the lysosome. there two possible causes. Firt , more diluted sample of lysosyme must be used .econd, Addition of Nacl or preincubation of sample with Nacl might be done. Nacl presence greatly reduces the active of lyosyme as well as EDTA . Thus , the turbidity will be less and absorbance of sample will be more.
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