The DelsaMax system Pre-Course Reading material DelsaMax PRO - PowerPoint PPT Presentation

BECKMAN COULTER INC. PARTICLE CHARACTERIZATION The DelsaMax system Pre-Course Reading material DelsaMax PRO DelsaMax Return to I ndex Service Training Press any key to continue Service Training Last update: July, 2013 Last update: May, 2013

Understanding Laser Light Scattering Theory Light scattering is a non ‐ invasive technique for characterizing macromolecules and a wide range of particles in solution . In contrast to most methods for characterization, it does not require outside calibration standards. In this sense it is an absolute technique. DelsaMax instruments make two different types of light scattering measurements for absolute molecular and particle characterization: • Classical Light Scattering / Static Light Scattering : Here, the intensity of the scattered light is measured as a function of angle. For the case of macromolecules, this is often called Rayleigh scattering and can yield the molar mass, rms radius, and second virial coefficient ( A 2 ). For certain classes of particles, classical light scattering can yield the size, shape, and structure. • Quasi-elastic Light Scattering (QELS) or Dynamic Light Scattering (DLS) : In a QELS measurement, time ‐ dependent fluctuations in the scattered light signal are measured using a fast photon counter. QELS measurements can determine the hydrodynamic radius of macromolecules or diameter of particles. DelsaMax Service Training Last update: July, 2013

The DelsaMax family The DelsaMax family is a premium nanoparticle characterization tool for the measurement of particle size and zeta potential. The DelsaMax family allows users to make measurements extremely fast with very low sample volumes (as low as 1 microliter volume in 1 second). DelsaMax PRO DelsaMax CORE DelsaMax ASSIST DelsaMax Service Training Last update: July, 2013

The DelsaMax PRO Light Scattering Analyzer The DelsaMax PRO combines a unique 31 detector diode array with an advanced Avalanche Photodiode Detector (APD) which enables analysis of both zeta potential and size virtually simultaneously. Each measurement takes approximately one second to complete. The optical design is patent pending. Key differentiators include optical design, easy injection port, and low sample volumes. The Pro is the top of the line, most capable instrument and includes both flow cells and cuvette based measurement options. DelsaMax PRO DelsaMax Service Training Last update: July, 2013

Intended Use / Overview Intended Use: The DelsaMax PRO is a bench top analyzer that provides non ‐ destructive electrophoretic mobility measurements and hydrodynamic radius of precious proteins, particles and macromolecules as small as 1 nm under dilute conditions, without perturbing these fragile species. Overview: The DelsaMax PRO incorporates an optical bench where the laser light source is split into a Sample Beam and a Reference Beam . The sample beam is scattered by the moving particles, collimated, and recombined with the reference beam onto an array of 30 photo detectors. Each of these photo detectors makes an independent measurement of the sample mobility in a process referred to as Massively Parallel Phase Analysis Light Scattering (MP ‐ PALS). These signals are then processed in real time with a proprietary demodulation algorithm. The Flow Cell has 1.6 mm spacing of the electrodes. This small gap reduces the required sample volume, decreases the required applied voltages, and further reduces potential sample damage. The simultaneous measurement of the hydrodynamic radius (for molecules) and diameter (for particles) are available with the embedded Quasi ‐ elastic light scattering (QELS). The integration of the QELS into the instrument utilizes proprietary optics, high quantum efficiency detection, and data processing to analyze backscattered light DelsaMax PRO DelsaMax Service Training Last update: July, 2013

DelsaMax PRO Optical Bench Block Diagram DelsaMax PRO DelsaMax Service Training Last update: July, 2013

Optical bench Laser assembly Optical Bench : Cell block The laser system, optics and piezo modulator, flow ‐ cell assembly, and multi ‐ channel photodiode array are all anchored to the base plate to provide a single, stable optical bench. The 50 mW diode pumped solid state (DPSS) single ‐ longitudinal ‐ mode narrow beam laser (diameter 0.08 mm) vertically polarized provides the light source for the system. Laser monitor signals are incorporated Piezo assembly into the laser assembly (Laser Power and Forward Monitor). The output of those monitors can be displayed on the Main panel on the instrument’s display . The Laser Power measures the intensity of the beam at the laser assembly before it enters the cell while the Forward Monitor measure the transmitted light through the sample cell and sample. The primary usage of the Forward Monitor is simply as a diagnostic tool to determine if there are bubbles or other foreign particles in the cell. PALS photo detectors assembly DelsaMax DelsaMax PRO Service Training Last update: July, 2013

Principles of Operation The DelsaMax PRO light scattering analyzer implements two separate optical systems based on:  PALS (Phase Analysis Light Scattering).  QELS (Quasi-Elastic Light Scattering). and can collect both QELS and PALS data at the same time on the same sample. DelsaMax PRO DelsaMax Service Training Last update: July, 2013

Principles of Operation Phase Analysis Light Scattering (PALS): The laser beam is split into a Sample Beam and a Reference Beam . The sample beam is scattered by the moving particles, is collimated, and then mixed/heterodyned with the reference beam directly onto an array of 31 photo detectors that make independent measurements of the electrophoretic mobility. This technique is called Massively ‐ Parallel Phase Analysis Light Scattering (MP ‐ PALS). The signals from the array elements are processed in real time with a proprietary demodulation algorithm. The detectors used in the PALS array are linear and robust over the dynamic range. The multiple detectors used in the DelsaMax PRO analyzer efficiently average away the effect of Brownian motion and reveal the electrophoretic velocity, even with low applied voltages. DelsaMax PRO DelsaMax Service Training Last update: July, 2013

Principles of Operation Dynamic Light Scattering (DLS): Sometimes referred to as Quasi ‐ Elastic Light Scattering (QELS), is a non ‐ invasive, well ‐ established technique based on Photon Correlation Spectroscopy (PCS) for measuring the size and size distribution of molecules and particles typically in the submicron region, and with the latest technology lower than 1nm. QELS is an internally installed function that measures time ‐ dependent fluctuations in the back scattered light signal using a fast photon counter. The fluctuations will occur rapidly for smaller, faster moving particles and more slowly for larger, slower moving particles. QELS measurements can determine the hydrodynamic radius of macromolecules or diameter of particles. The fluctuations of the scattered light are analyzed using the autocorrelation function. DelsaMax PRO DelsaMax Service Training Last update: July, 2013

Technical Specifications Measurement range: Size: 0.2 nm to 5000 µm, hydrodynamic radius Mobility: No practical limits Zeta Potential: No practical limits exist Cell Voltage range: 0 to 3V typical. Max 600 V/cm. < 5 x 10 7 Daltons (g/mol) Molecular weight: Light source: Laser Diode 532 nm (green), 50 mW DPSS (diode-pumped solid state) Laser classification: Class 1 for operation (no access to radiation) Class 3B for service and maintenance (trained personnel only). Laser lifetime: 10,000 hours Detection system: - PALS (Phase Analysis Light Scattering) photodiode array for multi- angle detection and mobility measurements. 31 detectors for analysis - 1 detector for Laser monitor. - Single photon counting module for QELS option. 1 detector . Instrument options: QELS (Embedded Quasi-Elastic Light Scattering) for size measurements. DelsaMax PRO DelsaMax Service Training Last update: July, 2013

Technical Specifications Scattering angle: Mobility (Zeta Potential): 4° to 15° (MP PALS) Hydrodynamic radius (Size): 171° (DLS - QELS) Correlator: Multi-Tao function by hardware for correlation curve Sample volume: Hydrodynamic radius (Size): 45 μ L Mobility (Zeta Potential): 170 μ L Sample concentration: Up to 40 % w/v Operating Temperature: 10°C to 35°C (50°F to 95°F), Analysis temperature range: 4°C to 70°C (39.2°F to 158°F), regulated by a Peltier. Power Supplies: +5V, +12V, ± 15V and +48V Power Voltage: 90 ~ 250 VAC. Universal Power Input. Frequency: 50 to 60 Hz inclusive. Consumption: 200 W (VA) typical. Fuses: Two fuses 4A 250V Slow Blow Dimensions: 59 cm (23”) x 36 cm (14”) x 21 cm (8”) Weight: 19 kg (41 lbs.) DelsaMax PRO DelsaMax Service Training Last update: July, 2013

Controls, Connectors & Indicators Fluid connectors cover Keypad Power On/Off switch LCD display Front view Fluid connectors cover: In/Out fluid connectors for plumbing the instrument are behind this cover. LCD display (touchscreen ): The LCD display allows you to monitor, control, and configure the DelsaMax PRO light scattering analyzer. Keypad: The keypad allows you to control the LCD display. On/Off switch: Main power on/off switch. DelsaMax PRO DelsaMax Service Training Last update: July, 2013

Controls, Connectors & Indicators Cell block Flow Cell Front view Cell block Cell block: Comes with a pop up door to allow access to the flow cell. DelsaMax PRO DelsaMax Service Training Last update: July, 2013