rev: July 22, 1999
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Extraction Protocol for IBL Steroid Elisa Tests code cat#RDI-RE52xxx
1) Material for Preparation of Tests:
a) Diethyl-ether
b) human serum free from steroid
c) 1.5ml safe-lock test tubes (eppendorf)
d) vortexing device VF2
e) centrifuge
f) -80 DEG C freezer
g) Sonorex ultrasound bath
h) pipets 40-2000ul
Extraction Procedure:
a) Mix 230ul serum with 1ml diethyl-ether
b) Vortex for 20 seconds and shake thoroughly for 15 minutes
c) Centrifuge for 5 minutes at >1500 g
d) Freeze-out upper organic phase at -80DEG C.
e) Repeat steps 1-4 and bring the extracts together
f) Evaoprate extracts down to 100ul (e.g. with nitrogen) in a water bath at 35 DEg C. Wash surface of the tube with 150ul of diethyl-ether, vortex shortly
g) Evaporate extracts down to about 20ul and add 75ul steroid free serum
h) Centrifuge for 3 minutes at >1500 X g
i) Evaporate the remaining diethyl-ether
j) Place the tube for 12 minutes in an ultra sonic bath
k) Centrifuge for 3 minutes at /1500 X g
l) The clear liquid sample may be used as described in the assay
procedure..
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