1.) In previous experiments, you determined that more than 90% of intact P falci
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Question
1.) In previous experiments, you determined that more than 90% of intact P falciparum cells and nuclei in a crude homogenate pellet in 10 minutes at 4500 rpm in a Sorvall GSA rotor. The GSA rotor has gone missing, so you borrow an SS-34 rotor from a neighboring lab. Predict how this difference in size and shape may affect the centrifugation parameters.
2.) Use the charts below to determine the appropriate speed and time for the Sorvall SS-34 rotor. Remember that your goal is to pellet intact cells and nuclei and leave the rest of the subcellular fragments in the supernatant.
(NOTE: In our textbook, it says that large structures (i.e. intact cells, nuclei, etc.) are separated at low speeds via differential centrifugation.)
Table 1-9. GSA Rotor: RCF and K Factor RCF (Radius) minimum r average Angular Velocity (rev/min) 9.27 cm 3.65 in) 11.94 cm 14.63cm (5.75 in) (4.70 in) 1 000 1 500 2 000 2 500 3 000 3 600 4 000 4 500 5 000 5 500 6 000 6 500 104 233 414 647 932 1 270 1 660 133 300 534 834 1 200 1 630 2 130 2 700 3 330 4 030 4 800 5 640 6 540 500 8 540 9 640 10 800 12 000 13 300 14 700 16 100 17 600 19 200 20 800 22 500 367 653 1 020 470 2 000 2 610 3 300 4 080 4 950 5 870 6 890 8 000 7 500 8 000 8 500 9 000 9 500 10 000 10 500 11 000 11 500 12 000 12 500 13 000 2 590 3 130 3 730 4 380 5 080 5 830 6 630 7 480 8 390 9 350 10 400 11 400 12 500 13 700 14 900 16 200 17 600 10 400 11 800 13 200 14 700 16 300 18 000 19 700 21 700 23 500 25 500 27 600Explanation / Answer
1- increase in the speed will affect the viability of cells. The GSA ROTORS has maximum RCF as 27,000 g and SS-34 has maximum RCF as 50,000 g. So even if u follow the same rpm the cells viability will vary as per the protocols you have used. Often increase in centrifugation speed will decrease cell viability.
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