Hemoglobin degradation in human erythrocytes with long-duration near-infrared laser exposure in Raman optical tweezers

Abstract

Near-infrared laser (785-nm)-excited Raman spectra from a red blood cell, optically trapped using the same laser beam, show significant changes as a function of trapping duration even at trapping power level of a few milliwatts. These changes in the Raman spectra and the bright-field images of the trapped cell, which show a gradual accumulation of the cell mass at the trap focus, suggest photoinduced aggregation of intracellular heme. The possible role of photoinduced protein denaturation and hemichrome formation in the observed aggregation of heme is discussed.

Raman Spectroscopy of Blood and Blood Components

“…[54][55][56][57] As LTRS technology has matured, it has been used to refine previous studies that had experienced limitations due to sampling protocols. Dasgupta 58 These bands were consistent with hemoglobin existing in an aggregated form, confirming earlier work by researchers from the Monash Figure 3. Raman spectra of oxygenated single erythrocytes recorded using different excitation wavelengths.…”

Long term Raman spectral study of power-dependent photodamage in red blood cells

“…Raman spectral dynamics showed significant and irreversible changes as a function of trapping duration that we attribute to a combination of photodamage of hemoglobin at short times followed by diffusion of hemoglobin out of the cell at longer times. Optical tweezers are receiving much attention in contemporary research, particularly in the biomedical sciences, [1][2][3][4][5][6][7][8][9][10][11][12][13] because the ability tweezers provide to manipulate individual cells with high precision. The use of a focused optical beam to immobilize cells without direct contact helps to avoid physical or chemical methods that often lead to undesirable surface-induced effects.…”

“…2,13 Optical tweezers have been widely used in association with Raman spectroscopy, in order to obtain accurate information about biochemical changes from a single cell. 5,8,9,11,14 However, to acquire high-quality Raman spectra of individual cells, irradiances larger than 1 GW/m 2 are typically used. In the absence of strong absorption, cells may withstand these exposures.…”

“…3,6,8,11,12 Some of these effects can be useful for monitoring cell function, as Liu et al have used laser-tweezers Raman spectroscopy to show that red blood cells reversibly deoxygenate when exposed to the forces within an optical trap, 11 and that the ability of cells to respond to optical forces can be used as a functional test for red blood cells. 12 The changes observed by Liu et al were shown to be reversible, however, Wood et al 3,7 and Dasgupta et al 8 have observed irreversible photodamage using laser tweezers Raman spectroscopy. Both of these studies attribute the observed Raman spectral changes in response to laser exposure as arising from photodegradation of hemoglobin, however, the two groups differ as to what photodamage is occurring.…”

“…Wood et al ascribe the spectral changes to methemoglobin formation and excitonic interactions induced by heme aggregation, 7 while Dasgupta et al assign the changes to the formation of hemichrome. 8 However, to date, there have not been any reports on the literature characterizing the temporal evolution of this damage at long time scales.…”