sdscomplexmix

Electrophoresis
Development & Consulting
Dr. Hanspeter Schickle
Wolfgang Gstrein

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complex samples

April 28, 2023

Complex samples such as E.coli dry powder - plant tissues - etc. tend to agglutinate during the stacking process due to the SDS-fat mycelles.
After this false starting process the sample form a “channel” in the track.
Remedy:
1. Use a extraction buffer without SDS.
This inhibits the solubilization of fat. Afterwards the fat-SDS-mycelles will disturb the electrophoresis.
The SDS-samples should be produced out of this concentrate.
2. Add 0.6 M Urea to the samples after the heating process.


	ad 1: Extraction Buffer (100 ml)

250 mM TRIS	3.04g
190 mM Glycine	1.5 g
1 mM EDTA	30 mg
0.01% Dodecylmaltoside	10 mg
0.005% Triton X100 (1%)	500 ul
(add freshly: 3 M Urea)	(180 mg / 1 ml)


	Fill up to 100 ml. The pH-value should be ~9.2


	ad 1: extraction procedure:

Ultrasonic treatment	15 min
Centrifuge 12000 rpm	5 min


	Then the SDS-sample procedure should follow


	ad 2: adding urea

	after the SDS-sample procedure: After the samples cooled down, add 3 M Urea (180 mg/1 ml) and shake well






	sample #4 is E.coli directly prepared with SDS sample-buffer.




	... samples #3-7 is E.coli prepared with extraction buffer


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