A standard method for preserving bone tissue samples is to embed them in paraffin or plastic (methyl methacrylate/dibutyl phthalate). Then they can be sectioned at 5-10 ?m thicknesses for transmission infrared experiments. These thin sections are required due to the extremely large signals provided by the phosphate and protein contributions to the infrared spectra. Although embedding provides an excellent means for examining the mineral components of bone, this technique has a few drawbacks including (1) interference of the infrared absorbance of the embedding medium, and (2) it is uncertain whether the embedding process affects protein secondary structure. Conventional freeze/microtome cleavage methods can be used to provide ?native? tissue samples. However, it is difficult to section to less than 20 ?m, which is problematic due to the high infrared absorbance at those thicknesses. Thus, we are using attenuated total reflection infrared microspectroscopy to examine such native samples (eg. without plastic). Sections of tissue are frozen and cleaved with a microtome and examined using Attenuated Total Reflectance (ATR). ATR is a surface technique that probes the tissue structure approximately 10 ?m into the surface based on the evanescent wavefront of the infrared radiation. Due to this property, the samples are examined ?wet?. ATR infrared microspectroscopy is a superior method for examining protein spectra, since no denaturing conditions that may perturb the protein structure are required.

National Institute of Health (NIH)
National Center for Research Resources (NCRR)
Biotechnology Resource Grants (P41)
Project #
Application #
Study Section
Project Start
Project End
Budget Start
Budget End
Support Year
Fiscal Year
Total Cost
Indirect Cost
Albert Einstein College of Medicine
United States
Zip Code
Vongsvivut, Jitraporn; Fernandez, Jason; Ekgasit, Sanong et al. (2004) Characterization of supported cylinder-planar germanium waveguide sensors with synchrotron infrared radiation. Appl Spectrosc 58:143-51
Masip, Lluis; Pan, Jonathan L; Haldar, Suranjana et al. (2004) An engineered pathway for the formation of protein disulfide bonds. Science 303:1185-9
Huang, Raymond Y; Miller, Lisa M; Carlson, Cathy S et al. (2003) In situ chemistry of osteoporosis revealed by synchrotron infrared microspectroscopy. Bone 33:514-21
Rashidzadeh, Hassan; Khrapunov, Sergei; Chance, Mark R et al. (2003) Solution structure and interdomain interactions of the Saccharomyces cerevisiae ""TATA binding protein"" (TBP) probed by radiolytic protein footprinting. Biochemistry 42:3655-65
Uchida, Takeshi; Takamoto, Keiji; He, Qin et al. (2003) Multiple monovalent ion-dependent pathways for the folding of the L-21 Tetrahymena thermophila ribozyme. J Mol Biol 328:463-78
Taylor, Colleen M; Watton, Stephen P; Bryngelson, Peter A et al. (2003) Inner-sphere complexation of cobalt(II) 2,9-dimethyl-1,10-phenanthroline ([Co(neo)]2+) with commercial and sol-gel derived silica gel surfaces. Inorg Chem 42:312-20
Dhavan, Gauri M; Crothers, Donald M; Chance, Mark R et al. (2002) Concerted binding and bending of DNA by Escherichia coli integration host factor. J Mol Biol 315:1027-37
Uchida, Takeshi; He, Qin; Ralston, Corie Y et al. (2002) Linkage of monovalent and divalent ion binding in the folding of the P4-P6 domain of the Tetrahymena ribozyme. Biochemistry 41:5799-806
Tang, Qun; Carrington, Paul E; Horng, Yih-Chern et al. (2002) X-ray absorption and resonance Raman studies of methyl-coenzyme M reductase indicating that ligand exchange and macrocycle reduction accompany reductive activation. J Am Chem Soc 124:13242-56
Guan, Jing-Qu; Vorobiev, Sergeui; Almo, Steven C et al. (2002) Mapping the G-actin binding surface of cofilin using synchrotron protein footprinting. Biochemistry 41:5765-75

Showing the most recent 10 out of 68 publications