High Z Metal Carbonyls for Imaging and Microspectroscopy

V. N. Joshi*, D. Ramamurthy*, R. D. Powell*, F. R. Furuya* and J. F. Hainfeld**

*Nanoprobes, Incorporated, 95 Horse Block Road, Yaphank, NY, 11980
**Biology Department, Brookhaven National Laboratory, Upton, NY 11973

Recently, we and others reported construction of electron density maps of tetra-iridium (Ir₄) cluster labeled virus capsids and ribosome particles by cryo-EM and X-ray crystallography respectively at medium to high resolution [1, 4]. These Ir₄ clusters contain carbonyl ligands that exhibit extremely intense infrared (IR) absorptions in the region where most of the biological materials do not absorb (Fig. 1b). This unique feature of metal carbonyls has led to the development of non-isotopic IR spectroscopic immunoassays [5]. We recently demonstrated that functionalized heavy atom carbonyl cluster labeled reagents can be used for bio-sensing/bio-recognition using Fourier transform infrared (FTIR) spectroscopy, albeit with lower detection sensitivities than competing technologies [6]. To demonstrate the use of heavy metal carbonyl clusters as dual labels for electron microscopy and FTIR microspectroscopy, we conjugated the previously reported Ir₄ label [4] to freshly reduced goat anti-rabbit F(ab) fragments following standard protocols [7]. The Ir₄ labels are clearly visible in dark-field scanning transmission electron micrograph (STEM) of the Ir₄-labeled F(ab) conjugate (Fig. 2a). The Ir₄-F(ab) conjugate was captured by 100 ng of covalently immobilized rabbit IgG on IR reflective slides following reported procedures [6,8]. The Ir₄ labels could be localized at 10x10 micron resolution by FTIR microspectroscopy (Fig. 2b).

Since IR microspectroscopic imaging has been used to distinguish chemical differences between healthy and diseased tissues [9], we envisage the use of metal carbonyl cluster labels for rapid localization of diseased tissues. Subtraction of the Ir₄ label spectrum from that of localized targets will decipher chemical information and underlying chemistry which other methods, such as light or fluorescence microscopy, do not. The Ir₄ labels reported here have active Raman vibrations. Since Raman microspectroscopy has also been used to distinguishing healthy and diseased tissues, we are investigating the use of metal carbonyls as labels for Raman microspectroscopy.

References

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Fig. 1. Schematic diagram of the Ir4 cluster label (a) and its IR spectrum (b).

Fig. 2. a) STEM of Ir4-F(ab) (full image width is 128 nm), and b) IR image of Ir4-F(ab) captured by immobilized IgG (calculated as peak height ratio of 1990/2150 cm-1; the blue colored regions indicate highest concentration).

© 2005 Cambridge University Press. Used with permission.