1.3

Corrections

Read the plate only once

Pipet 150 µL of each unknown in E3 - 12 as needed

Programs

• Serial2&5_Abs
• Serial2&5_FL
• E1 &E2: blank E3-E12 for unknowns
• F1 - F12: dilute by 2
• G1 - G12: dilute by 5

Materials:

• ~1 mL 1mM fluorescein (try to make it right via Abs)
  • E = 50358/M
  • A = 50358/M * C
  • at 10 µM, A = 0.5038
  • dilute 1 mM 1/100 to make 10 µM, take Abs
• 5 mL 10 mM NaOH
• black plate from last time (or new one if old one has no clean wells)

Conversion factors

Abs = 59.7*(mM)-0.341 up to mM=0.03125

FL = 10^6 * mM +173.06 up to mM = 0.008

FL = 2 X 10^6 * mM + 19.624 up to mM = 0.00195

Unknowns

• Make dilution series from the "1mM" stock.
• find lowest useful concentration for Abs. Dilute slightly for unknown 1 - must be done with FL. Abs is bad below 0.0005 and meaningless below 5 x 10^-7mM
• find a concentration too high for fluorescence, but OK for abs, e.g., 0.05.
• find a concentration too high for either. must be >0.125

2013: