Staining, Histochemistry and Histotechnology
(Frequently Asked Questions)
Dr. John A. Kiernan
Department of Anatomy
and Cell Biology
The University of Western Ontario
Carbodiimides as fixatives
Does anyone know what carbodiimide is and how it works?
The name "carbodiimide" is sometimes applied to cyanamide (hydrogen cyanamide, H2NCN. Don't confuse this with calcium cyanamide, CaNCN.), which does not seem to have been used as
Carbodiimides are compounds that combine with and cross-link carboxyl groups. They fix proteins by joining together C-termini and/or side chains of glutamic and aspartic acid units. Their general chemical formula is R-N=C=N-R'
In contrast, aldehydes combine mainly with protein nitrogen atoms. Cross-links between the lysine side-chain amino group and the amide nitrogens of peptide linkages are thought to do most of the fixing.
Various carbodiimides have been used as fixatives over the
years, but they have never caught on in a big way. They are the sort of things used when more ordinary compounds are unsuitable. See Pearse's Histochemistry, Vol. 1 (3rd ed., Churchill-Livingstone, Edinburgh, 1980) page 107 for a proper account.
If the antigenicity of a protein is critically dependent on free amino groups of an epitope, then one of the carbodiimide fixatives might be a sensible alternative to formaldehyde. If it's for paraffin sections, a chemically unreactive fixative such as Clarke's or Carnoy's might be even more sensible.
John A. Kiernan,
Department of Anatomy & Cell Biology,
The University of Western Ontario,
LONDON, Canada N6A 5C1