Fong:Spectrophotometry

<owwmenu font="arial, helvetica, sans-serif" bold="1"

       color="White" bgcolor="gray" hovercolor=#FF9933
       bghovercolor="black" topfontsize="7" fontSize="7" pagewidth="750"
       image="Cth.JPG" lab="Fong">
   Home=
   Lab Members=People
   Research=Research
   Internal=Internal, Protocols=Protocols, Chemical Inventory=Inventory
   Links=Links

</owwmenu>

Overview

Materials

• Blank
• Sample
• Fingers
• Spectrophotometer
• An expensive and fragile cuvette

Procedure

You need at least 120 μL for a good reading; dilute into water appropriately (eg. 20x dilution is 6 μL sample in 114 μL water). Also if you are reading DNA or other sample with a buffer, use a similar buffer at the same concentration in preparing your blank.

1. If the spec's in lamp-off mode, press any button with your finger.
2. Press "Esc" until you reach the main menu.
3. Select the type of reading you are looking for.
4. Enter the correct λ for your sample
5. Add at least 120 μL of your prepared blank to the cuvette.
6. Measure the blank and thoroughly rinse out the cuvette.
7. Add at least 120 μL of your sample to the cuvette.
8. Use the measured absorbance to calculate concentration.
9. ?
10. Profit.

Calculation

Take your absorbance at λ and multiply it by the dilution factor and the constant for the sample in question (e.g., C_DNA = 50) The number obtained is the concentration in μg/mL.