INSTITUTE OF PHYSICS PUBLISHING PHYSICS IN MEDICINE AND BIOLOGY Phys. Med. Biol. 48 (2003) 243–257 PII: S0031-9155(03)39902-6 Methods for measuring the infrared spectra of biological cells Judith R Mourant, Rowena R Gibson, Tamara M Johnson, Susan Carpenter, Kurt W Short, Yujiro R Yamada and James P Freyer Bioscience Division, MS E535, Los Alamos National Laboratory, Los Alamos, NM 87545, USA Received 23 July 2002, in final form 27 November 2002 Published 8 January 2003 Online at stacks.iop.org/PMB/48/243 Abstract Infrared (IR) spectroscopy of biological cells is a growing area of research, with many papers focusing on differences between the spectra of cancerous and noncancerous cells. Much of this research has been performed using a monolayer of dehydrated cells. We posit that the use of monolayers can introduce artefacts that lead to an apparent but inaccurate measurement of differences between cancerous and noncancerous cells. Additionally, the use of dried cells complicates the extraction of biochemical information from the IR spectra. We demonstrate that using suspensions of viable cells in aqueous suspension reduces measurement artefacts and facilitates determining the concentration of the major biochemical components via a linear least- squares fit of the component spectra to the spectrum of the cells. 1. Introduction Many of the reported measurements of infrared (IR) spectra have been performed on dried cells (Boydston-White et al 1999, Ramesh et al 2001, Cohenford and Rigas 1998). The IR spectra of dried cells may differ from the IR spectra of cells in their natural aqueous state since the spectra of many of the known biochemical components of cells change with hydration level. The IR spectra of proteins have been shown to change dramatically after dehydration (Dong et al 1996, Pevsner and Diem 2001a, Prestrelski et al 1993, Jackson and Mantsch 1995, van de Weert et al 2001). Absorption bands can shift and change in amplitude as a function of hydration. The spectra of nucleic acids also depend on hydration status. As RNA and DNA are hydrated, the phosphate absorption bands narrow (Pevsner and Diem 2001b) and the conformation of DNA can be altered (Kim et al 1997). In addition to spectral artefacts caused by dehydration of the sample, another potential source of spectral distortions is inhomogeneity of the sample. As presented in the results section, the absorption spectrum of a material can differ depending on whether the sample is homogeneous or heterogeneous. Three potential causes of sample inhomogeneity are 0031-9155/03/020243+15$30.00 © 2003 IOP Publishing Ltd Printed in the UK 243