Need help with the highlighted calculation\'s. (there are two in total) Will giv

Question

Need help with the highlighted calculation's. (there are two in total) Will give a thumbs up.

Probing Blot with anti-GFP primary antibody 8. Calculate how much primary antibody you need to add to 5.0 ml of TBST buffer to give a 1:3333 dilution of the primary antibody (mouse anti-GFP.) CALCULATION You will incubate your blot with primary antibody with another group (meaning 2 blots in one tube/tray at the same time with twice the volume of TBST+ primary antibody) So, how much primary antibody you need to add to 10ml of TBST buffer to give a 1:3333 dilution of the primary antibody? AETLO One of the two groups should use the 5ml pipet at the bench to transfer 10.0 ml of TBST buffer into one of the "Falcon" conical tubes. IMPORTANT: You will use the 10 ml pipet for your cells in protocol #2) 9. 10. Bring this to your TA, who will pipette the calculated amount of antibody into the 10ml for you. 11. Pour the diluted antibody/TBST solution into one tray and add both blots to the tray. Incubate the blots on an orbital shaker at room temperature for 1 hour. Flip the position of the blots (which is on the top ys bottom) half-way through the incubation.

Explanation / Answer

8. primary antibody dilution required = 5000 ?l/ 3333 = 1.5 ?l

14. Secondary antibody dilution required = 5000 ?l / 667 = 7.5 ?l ( approx)

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