Volume 87, Issue 2, Pages (August 2004)

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Presentation transcript:

Volume 87, Issue 2, Pages 1298-1306 (August 2004) Elastic Light Scattering from Single Cells: Orientational Dynamics in Optical Trap  Dakota Watson, Norbert Hagen, Jonathan Diver, Philippe Marchand, Mirianas Chachisvilis  Biophysical Journal  Volume 87, Issue 2, Pages 1298-1306 (August 2004) DOI: 10.1529/biophysj.104.042135 Copyright © 2004 The Biophysical Society Terms and Conditions

Figure 1 Diagram of single-beam optical trap used for levitation of a single particle. Biophysical Journal 2004 87, 1298-1306DOI: (10.1529/biophysj.104.042135) Copyright © 2004 The Biophysical Society Terms and Conditions

Figure 2 Schematics of optical setup used to record scattering diagrams from single particles suspended in a optical trap. Biophysical Journal 2004 87, 1298-1306DOI: (10.1529/biophysj.104.042135) Copyright © 2004 The Biophysical Society Terms and Conditions

Figure 3 Comparison between experimental scattering diagram for 5-μm PMMA bead and Lorenz-Mie theory light-scattering calculation. Calculation parameters: relative refraction index nrel=1.122; particle radius, 2.385μm; polarization, linear, azimuthal angle, 45° (defined as an angle between the polarization of the laser and detection plane). Biophysical Journal 2004 87, 1298-1306DOI: (10.1529/biophysj.104.042135) Copyright © 2004 The Biophysical Society Terms and Conditions

Figure 4 Images of a trapped A375 cell and a 6.2-μm Silica bead at two different time points as viewed from the bottom of the cylindrical cuvette. Direction of incident excitation laser light is right to left as shown. The images on the right were recorded ∼3s later than images on the left. The particles were located in the beam waist of the excitation laser. Biophysical Journal 2004 87, 1298-1306DOI: (10.1529/biophysj.104.042135) Copyright © 2004 The Biophysical Society Terms and Conditions

Figure 5 Scattering diagram of a single A375 cell suspended in a optical trap. The data shown are after subtraction of the background signal. Biophysical Journal 2004 87, 1298-1306DOI: (10.1529/biophysj.104.042135) Copyright © 2004 The Biophysical Society Terms and Conditions

Figure 6 Time-dependent scattering diagrams from an A375 cell shown for two different timescales. An important feature seen in both plots is that the forward scatter region is much more stable than the side-scatter region. In A, intensity fluctuations are already quite noticeable in the 40-ms time steps, specifically in the 90° region. Note that the intensity is plotted on a log scale, so that small differences are actually quite large. The plot shown in B fluctuates in both peak values and angular position of peaks. Notice the region near 45°, where a peak actually turns into valley, or rather the peak has shifted to the right. This variation in the scatter pattern was evident in all trapped cells. Biophysical Journal 2004 87, 1298-1306DOI: (10.1529/biophysj.104.042135) Copyright © 2004 The Biophysical Society Terms and Conditions

Figure 7 Time-dependent integrated light-scattering signals at forward (A) and side (B) scatter angles and axial optical force (C) measured for a A375 cell suspended in optical trap. Cell is released from the trap at time T=2.5s. Biophysical Journal 2004 87, 1298-1306DOI: (10.1529/biophysj.104.042135) Copyright © 2004 The Biophysical Society Terms and Conditions

Figure 8 Forward scatter and axial optical force from a suspended A375 cell as function of time. Biophysical Journal 2004 87, 1298-1306DOI: (10.1529/biophysj.104.042135) Copyright © 2004 The Biophysical Society Terms and Conditions

Figure 9 Time-dependent light-scattering signal integrated in four progressively larger angular ranges. Biophysical Journal 2004 87, 1298-1306DOI: (10.1529/biophysj.104.042135) Copyright © 2004 The Biophysical Society Terms and Conditions

Figure 10 Time-dependent integrated light-scattering signals (at forward and side-scatter angles) and axial optical force measured for a 6.2-μm bead suspended in optical trap. Biophysical Journal 2004 87, 1298-1306DOI: (10.1529/biophysj.104.042135) Copyright © 2004 The Biophysical Society Terms and Conditions

Figure 11 Time dependence of side-scattering signal for a A375 cell and 6.2-um Silica bead (upper graph) and corresponding Fourier amplitude spectra (lower graph). Biophysical Journal 2004 87, 1298-1306DOI: (10.1529/biophysj.104.042135) Copyright © 2004 The Biophysical Society Terms and Conditions

Figure 12 Light-scattering diagrams of a normal blood lymphocyte and two different granulocytes. Biophysical Journal 2004 87, 1298-1306DOI: (10.1529/biophysj.104.042135) Copyright © 2004 The Biophysical Society Terms and Conditions