Abdullah MUT 20503836 03.12.

2014 Detection with Quadrant Photodetectors

I. Introduction Optical tweezers are proven to be effective in many areas since Arthur Ashkin show that they can be used to manipulate small particles even bacteria and viruses with non-contact approaches. [1,2] Due to the range of optical tweezers which lies around piconewtons and femtonewtons, they can be used both as a manipulator and measurement system. Basically, an optical tweezers setup is composed of a laser which can be used as both trapping light source and probing, several lenses for making use of the light at maximum efficiency, a stage for the sample, and finally a CCD camera and a quadrant photodiode for data acquisition. There can be changes like using two lasers, one for trapping and one for probing. Here the basic working principle of a quadrant photodiode will be explained, the usage of quadrant photodetectors for position measurement tool and hence a force measurement tool will be investigated and finally several applications that are using these devices will be presented briefly. II. Quadrant Photodetectors As the name gives out, a quadrant photodetector is a specially designed photodetector that is composed of four different detection sites that work separately for generating voltage as a result of the light that falls on them. The voltage contributions from different sites of the detector will change due to the position of the trapped particle. By the calibration of voltage generation and contribution to this voltage from these individual quadrant sites this device can be used as an effective tool for position and hence force sensing. III. Quadrant Photodetectors Calibration Since quadrant photodetectors are generating voltage as the output calibration of these photodiodes plays a crucial role for making a measurement with this tool. Without the calibration the output voltage does not mean anything or mean everything. In order to calibrate a quadrant photodiode, light from a trapped particle can be used since we are interested in optical tweezers. To start with there should be a known displacement scale for the calibration. The trapped particle should be moved with optical tweezers on this scale and the output voltage should be measured. This should be done both for x and y directions in order to gather information about the displacement on a plane. By doing this the quadrant photodetector can be used to track the position of the photodiode. After displacement calibration the quadrant photodetector is ready for force calibration in order to be used as a force measurement tool. In order to use a trapped particle as a photonic force microscopy (PFM) probe, the force on this probe should be measurable just like the force on the tip of an atomic force microscope. To calibrate the quadrant photodetector fluid dynamics is used. The force from a fluid, with known viscosity () and velocity (), exerted on a particle with radius (R) is given by the Stokes law.

When the force on the calibration particle from the fluid is equal to the trapping force of the optical tweezers the particle will stay stationary. By tuning the velocity of the fluid the displacement characterization of the trapped particle can be made. For a certain value of velocity the particle will no longer be stationary. By using this velocity the force on the particle can be calculated and hence the trapping force can be measured. This process is similar to pull a string. The displacement will increase

Abdullah MUT 20503836 03.12.2014 due to the increase in velocity and similarly increase in force. After a certain point displacement makes a jump since it is released for the trap. [3] So far the quadrant photodetector is calibrated for displacement and by the use of this displacement calibration, it is calibrated for force measurement. IV. Applications Since the optical tweezers is a non-contact method and the forces that can be measured are on the order of pico- femtonewtons, this method is highly useful in biology applications. The possibility of measurements of forces this small makes optical tweezers a useful tool for measuring the binding forces between biological structures. The first application is the measurement of a molecule, namely the A1 domain of von Willebrand factor, to glycoprotein Ib-IX complex of the cell membrane. This binding is important because it is the first stage of hemostasis and thrombosis. [4] For calibration of the quadrant photodetector a modified version of the Stokes law is used;

Where h is the distance between the center of the bead and the coverslip. Due to this force function they generate a trapping force versus quadrant photodetector output voltage graph. In order to measure the force between the A1 molecule and the GP Ib-IX, a 2 m diameter polystyrene bead is used as the A1 carrier. Then this optically trapped bead is moved towards a cell membrane that has GP Ib-IX on it. The A1 carrying polystyrene bead touches the membrane and then is detached slowly from the membrane by the optical tweezers. Due to the binding force, the bead is not moving at the beginning since the trapping force is not enough to break the bond. When the force is enough the bead detaches from the cell membrane in falls into the center of the optical trap where the center of the quadrant photodetector is set. As soon as the bead is separated from the cell membrane a drop in the displacement reading occurs. By this way the binding force between the molecule and the cell membrane is measured as around 13 pN. Another application similar to the one above measures the force between bacteria and protein coated surfaces. [5] The Stokes law used to calibrate the quadrant photodetector is same with the above except the numerator is half of it. In this case 10 m diameter polystyrene beads are used as the protein carrier. These beads are trapped and brought in contact to the bacteria. Then they are detached and the force is measured for full detachment. This detachment is done for different pulling rates. Different pulling rates seem to give different binding forces. For 1 m/s pulling rate the binding force is 50 pN although the binding force for 10 m/s is 60 pN. A similar work for optical tweezers perspective combines the optical trapping with a patch clamp. Patch clamp is a mechanism that holds the target cell. In this work the strength of the tethers is investigated. Tethers are rope like structures between a molecule and the cell membrane. The quadrant photodetector is calibrated according to the Stokes law. The trapped beads are brought close enough to cell membranes and after tether formation they are separated and the force is measured. In this example the patch clamp seems to change the force output voltage graphs. Since the patch clamp holds the cell steady, a saturated behavior is gathered for force. [6]

Abdullah MUT 20503836 03.12.2014 The quadrant photodetector calibrations until now are used the linear part of the calibration curves. However, it is shown that by the usage of the whole displacement versus output voltage curve and the cross-talk between different the output signals can be used to increase the detection range almost 10 fold with a 300 nm radius polystyrene bead. [7] V. Conclusions Quadrant photodetectors are proven to be useful and practical in PFM since their calibration procedure is well known. [3] The data acquisition is fast due to the camera based measurement methods. [7] As can be seen above it is also very open for manipulations. VI. References [1] Ashkin A. and Dziedzic J. M. ,Science,235,1517,1987 [2] Ashkin A. et. al., Nature,330,769,1987 [3] Hong-Lian, GUO et.al., Chin. Phys. Lett., 20, 6, 950, 2003 [4] Maneesh Arya ; Gabriel M. Romo ; Jose A. Lopez ; Bahman Anvari; Dynamic measurements of forces between thrombus-inducing proteins using optical tweezers. Proc. SPIE 4962, Manipulation and Analysis of Biomolecules, Cells, and Tissues, 224 (June 19, 2003) [5] Kathryn H. Simpson ; Gabriela Bowden ; Magnus Hook ; Bahman Anvari; Optical measurements of dynamic adhesive forces between bacteria and protein-coated surfaces. Proc. SPIE 4962, Manipulation and Analysis of Biomolecules, Cells, and Tissues, 263 [6] Feng Qian ; Sergey A. Ermilov ; David R. Murdock ; William E. Brownell ; Bahman Anvari; Membrane tether formation from voltage-clamped outer hair cells using optical tweezers. Proc. SPIE 5331, Nanobiophotonics and Biomedical Applications, 112 (June 1, 2004); [7] Perrone, S., Volpe G., and Petrov, D., Review Of Scientific Instruments, 79, 106101, 2008