Ca specific electrode, Ca chelators BAPTA or APTRA

Valery F. Thompson valeryt at ag.arizona.edu
Wed Mar 29 16:57:07 EST 1995


I am interested in using a Ca2+ specific electrode for measurement of free 
Ca2+ in solutions.  I have read that it is possible to measure down to 1µM 
Ca2+.  I am primarily interested in the range of 1-1000µM.  How reliable are 
the values at the 1µM level?  Is it possible to go lower?

I am also interested in hearing about possible interferences with the 
measurements.  I know Tris interferes, but at what level?  

I have tried using some of the chelators such as 4F- APTRA and 5,5'-dibromo 
BAPTA, but without much success.   The absorbance of 
these chelators does not seem to change very much (only about 0.02AU) when 
going from "Ca free" to "Ca satd" at a chelator concentration that would be 
acceptable for my experiments (about 25µM).  The purpose is to measure the 
free Ca conc in a protein solution, and 25µM of my protein is about 
2.5mg/ml (I am interested in the Ca binding properties of the protein).  
Unfortunately I only have about 10mg of it and it is very difficult to purify. 
Does anyone know how stable these chelators are in solution?

I have soaked all of my glassware and plasticware (except pipet tips and 
microcentrifuge tubes) in 10% HNO3 and rinsed them with high purity water.  I 
have previously measured the Ca contamination of my stock reagents by AA and 
found them to be below the detection limit.

If anyone has some information on the Ca specific electrode (I have requested 
literature from several companies, but haven't received it yet) or on the use 
of the chelators 5,5'-dibromo BAPTA or 4F-APTRA, I would appreciate hearing 
about it. 

Thanks.

Valery Thompson
Muscle Biology Group
University of Arizona
Tucson, AZ  85721
valeryt at ag.arizona.edu 



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