Advances in particle concentration measurements

Transcription

1 Advances in particle concentration measurements Dr Hanna Jankevics Jones Principal Applications Scientist

2 Company Confidential Outline Advances in particle concentration from Malvern Panalytical Capture Tracking Analysis Nanoparticle tracking analysis quick overview New! NanoSight Sample Assistant increase throughput and remove operator variation New! Multi-angle dynamic light scattering gives higher resolution and individual particle populations concentration 2 Title of the presentation

3 Company Confidential Visualisation Nanoparticle tracking analysis (NTA) Capture Tracking Analysis No need to know sample refractive index or density 3 Nanoparticle Tracking Analysis (NTA) 31/1/18

4 Size Measurement NanoSight tracks nanoparticles moving under Brownian Motion By tracking the particles, we can determine the diffusion coefficient and use it to calculate the size (Stokes-Einstein equation) Smaller particles move faster than larger particles ISO19430: Particle Tracking Analysis (PTA) method describes limitations, quantification parameters as well as instructions on how to operate the equipment in a certified manner. Capture Tracking Analysis 4 Nanoparticle Tracking Analysis (NTA) 31/1/18

5 Polydisperse Samples Sample distribution examples Monodisperse / bimodal broad, tails Multimodal: 100 nm, 200 nm, 300 nm, 400 nm PS latex Broad, tails, outliers, aggregation 5 Nanoparticle Tracking Analysis (NTA) 31/1/18

6 Concentration Concentration upgrade - can be done by the user in their lab NanoSight track individual particles, allowing to have their exact number The volume is determined by the field of view (X and Y) and from the laser beam profile (Z) Concentration upgrade for NS300 and NS500, *requires syringe pump 6 Nanoparticle Tracking Analysis (NTA) 31/1/18

7 Identify Subpopulation Relative refractive index or fluorescence Unknown mixture of gold and silica Smaller gold scatters more light than bigger silica Different laser wavelengths for different fluorophores (430) nm (500) nm (565) nm (650) nm 7 Nanoparticle Tracking Analysis (NTA) 31/1/18

8 NanoSight Sample Assistant 8 Title of the presentation

9 Samples Requirements and Compatibility 1ml per sample, using deep well plate Sample in water or aqueous buffer, using low volume flow cell (LVFC) Examples: Exosomes Viruses Low concentration proteins Drug delivery nanoparticles Metal nanoparticles Water treatment samples 9 Customer Intro NS Sample Assistant

10 M o d a l S iz e (n m ) C o n c e n tra tio n (p a r tic le s /m L ) Data repeatability 100nm polystyrene latex 96 well plate 3x60 second captures in flow-mode, data generated in 24 hours liposomes C o n c L 1 b u ffe r 1 L 2 b u ffe r 1 L 2 B u ffe r C o p y o f S iz e W e ll N u m b e r Size and concentration monitored over 15hours. % Coefficient of Variation Size (Mode, nm) Concentration (particles/ml) Expert user Sample COMPANY Assistant CONFIDENTIAL W e ll N u m b e r Most stable condition for these liposomes identified as L2 Buffer 2.

11 Summary NanoSight Sample Assistant Specifications Sample capacity System set-up time Sample measurement time 96 max (1 x 96 Deep well plates) Under 30 minutes Under 10 min/sample for a 3x60s measurement including clean and analysis Cross contamination <0.1% Required sample volume per sample Sample compatibility Availability 1000uL Aqueous (Low Volume Flow Cell) With new NS300 orders or as an After Sales accessory 11 Customer Intro NS Sample Assistant

12 Concentration and higher resolution size distribution from multi-angle dynamic light scattering Title 12 of the presentation

13 Higher resolution sizing and particle concentration per population by multi-angle dynamic light scattering What does it do? Combines scattering from multiple angles into one result giving sample size distribution and concentration What do we need? Scattering data from multiple angles Buffer scattering background data Material and dispersant absorption and refractive index System scattering sensitivity (one off reference sample measurement, normalising the attenuators) Laser 13 What does it give you? Cuvette based measurements of size and concentration for a range of materials: liposomes, gold, silica, protein, virus etc increased size resolution down to 1:2 for some systems* (e.g 150nm vs 300nm PS latex) Particle concentration (#/ml) for each resolvable particle Title of the presentation population

14 H y d r o d y n a m ic s iz e (n m ) In te n s ity (% ) concentration (mg/ml) Concentration measurements of 30nm gold particles collaboration with Caterina Minelli and Magdalena Wywijas 30 nm gold particles from BBI solutions 1.00E+12 y = 1E+11x R² = E E E m u lti a n g le D L S p e a k n m E Dilution factor D ilu tio n fa c to r H y d ro d y n a m ic d ia m e te r (n m ) 14 Title of the presentation

15 n o rm a lis e d c o n c e n tra tio n normalised concentration H y d r o d y n a m ic s iz e (n m ) Concentration repeatability Dependency on size measurement m u lti a n g le D L S p e a k n m 2.0 <40% RSD 19-28% RSD <13% RSD D ilu tio n fa c to r Scattering cross section dependency on size concentration normalised to mean D ilu tio n fa c to r normalised size 15 Title of the presentation

16 In te n s ity (% ) p e a k s iz e (n m ) In te n s ity (% ) C o n c e n ta tio n (# /m L ) Concentration from multi angle dynamic light scattering (#/ml) How accurate and repeatable are the concentrations? collaboration with Caterina Minelli and Magdalena Wywijas Fresh aliquot every morning from the same stock solution (kept in fridge) R e p e a t m e a s u r e m e n ts d a y n m g o ld fr e s h a liq u o t e v e r y d a y st n d 3 rd 4th 5th UV-Vis vs Multi-angle dynamic light scattering 1.00E E+11 y = 6.122x R² = H y d ro d y n a m ic d ia m e te r (n m ) 3 0 n m g o ld D a y 1.00E E D a y 1 D a y 2 D a y E E E E E E+12 Concentration from UV-Vis (#/ml) 1 0 D a y 4 D a y h y d ro d y n a m ic d ia m e te r [n m ] Title of the presentation D a y

17 Hydrodynamic diameter (nm) In te n s ity (% ) C o n c e n tr a tio n (# /m L ) Concentration (particles/ml) Liposome size and concentration measurements Examples from a dilution series from stock (36.5mg/mL HSPC/CHOL 55:45 Liposomes) down to 1:10,000, E : d ilu tio n *10^8 #/ml 1.00E+12 multi-angle DLS conc E+11 NTA *10^4 #/ml 1.00E E+09 H y d ro d y n a m ic d ia m e te r (n m ) H y d ro d y n a m ic d ia m e te r (n m ) 1.00E+08 NTA Total conc: 7.78*10^8 #/ml NTA - concentration Multi-angle DLS - concentration 1.00E+07 Too high conc Dilution factor Too low conc Dilution factor Title of the presentation 17

18 Summary Advances in particle concentration measurements NTA and Sample Assistant provides higher throughput NTA measurement, with reduced variation Cleaning protocols ensure reliable sampling and minimises crosscontamination Sizing, polydispersity, sub-population separation and concentration all available with the Sample Assistant Flexible software to set up multiple different measurements across a plate, to reduce operator time spent on measurements and analysis 18 Title of the presentation Multi-angle dynamic light scattering provides a higher resolution and concentration per population in a quick cuvette based measurement without the need of calibration or separation It can go down to 1:2 in resolution, e.g. 150 vs 300nm latex It can measure a wide range of sample materials such as gold, silica, liposomes etc The concentration range is across approx. 4-7 orders of magnitude depending on particle size and particle material

19 Thank you for your attention Questions? 19 Title Malvern of the Panalytical presentation 2018