Stretching Single DNA Molecules and using High-Speed Camera Power Spectral Analysis to Demonstrate High Force Capabilities of Holographic Optical Tweezers

Abstract

Holographic optical tweezers (HOTs), in which a Spatial Light Modulator is used to change the phase pattern of the laser light, enable the manipulation in three dimensions of many particles simultaneously. This can be used for the probing of extended structures such as cells and extended protein networks. To allow for quantitative force measurements, the HOT traps need to be calibrated. However, nanometer-scale position modulations are introduced by the Spatial Light Modulator. In power spectral analysis, modulations at specific frequencies and drift can be readily identified in the spectrum and omitted before analysis, making this the preferred method of calibration for our HOTs. We use high-speed camera imaging for position detection of multiple trapped particles simultaneously, from which we obtain power spectra with 1.25 kHz bandwidth. For stiff traps, however, blur due to image integration time affects the detected particle positions significantly. Taking the effects of blur, aliasing and position detection error into account, as put forward by Wong and Halvorsen [Opt. Express 14, 12517, 2006], we are able to obtain the corner frequency fc of the power spectrum for stiff traps with fc up to 3.5 kHz. We demonstrate the utility of our calibration approach by measuring the force-extension curve for 4-micrometer-long DNA.