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how to determine the chlorophyll content of chloroplasts using a spectrophotometric assay

来源:本站原创  2006/7/27 10:33:00   访问量:2387
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Overview
This protocol describes how to determine the chlorophyll content of chloroplasts using a spectrophotometric assay.
 
Procedure
1. Isolate Chloroplasts from spinach leaves (See protocol for preparation of chloroplasts from spinach leaves).

2. Pipet 1 ml of a chloroplast suspension into a 15 ml centrifuge tube and add 8 ml of Acetone. Mix thoroughly.

3. Add 1 ml of ddH2O, mix again and centrifuge at 1,000 X g for 3 to 5 minutes.

4. Measure the absorbance of the sample from Step 3 on a spectrophotometer at 652 nm (See Hint #1).

5. Calculate the amount of total chlorophyll in 1 ml of the Chloroplast suspension. The amount of chlorophyll is given by the equation:

mg chlorophyll per ml = Absorbance / 34.5

 
BioReagents and Chemicals
Acetone
 
Protocol Hints 
1. Use water as a blank to zero the spectrophotometer.

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