Available in two formats, Asymmetric Flow Field-Flow Fractionation (AF4) and Hollow-Fiber Flow Field-Flow Fractionation (HF5) provide numerous advantages over other separation techniques:
- Fractionate macromolecules and nanoparticles 1 nm to > 1 µm
- Eliminate column shear that may degrade fragile samples
- Avoid adverse interactions with column packing beads
Coupling an FFF system to a set of Wyatt MALS and/or DLS detectors creates a powerful system for accurate and robust characterization of molar mass and size distributions for simple or complex samples:
- Liposomes, virus-like particles or inactive viruses
- Whole serum
- Nanoparticles and biopolymers in the food industry
- Environmental nanoparticles
- Heterogeneous biopolymers and synthetic polymers
- Oligomers and aggregates
- Conjugates such as ADCs, PEGylated proteins or block co-polymers
Andres-Brull, M.; Al-Assaf, S.; Phillips, G. O.; Jackson, K. Optimisation of asymmetrical flow-field fractionation for the characterization of gum arabic (Acacia sengal var senegal) and comparison with gel permeation chromatography. Anal. Methods 2013, 5, 4047-4052.
Augsten, C.; Mäder, K. Characterizing molar mass distributions and molecule structures of different chitosans using asymmetrical flow field-flow fractionation combined with multi-angle light scattering. Int. J. Pharm. 2008, 351, 23-30.
Bousse, T.; Shore, D. A.; Goldsmith, C. S.; Hossain, M .J.; Jang, Y.; Davis, C. T.; Donis, R. O.; Stevens, J. Quantitation of influenza virus using field-flow fractionation and multi-angle light scattering for quantifying influenza A particles. J. Virol. Methods 2013, 193, 589-596.
Hupfeld, S.; Moen, H. H.; Ausbacher, D.; Haas, H.; Brandl, M. Liposome fractionation and size analysis by asymmetrical flow field-flow fractionation/multi-angle light scattering: influence of ionic strength and osmotic pressure of the carrier liquid. Chem. Phys. Lipids 2010, 163, 141-147.
Wyatt, P. Submicrometer particle sizing by multiangle light scattering following fractionation. J. Colloid Interf. Sci. 1998, 197, 9-20.
Instrumentation for FFF-MALS
Field-Flow Fractionation Systems
Asymmetric Flow Field-Flow Fractionation (AF4) and Hollow-Fiber Flow Field-Flow Fractionation (HF5) perform versatile separation of macromolecules and nanoparticles ranging from 1 nm to 10,000 nm in radius. Combined with MALS and DLS instrumentation, FFF separation is the ideal means for obtaining distributions of molar mass and molecular size, in solution.
Wyatt offers complete FFF-MALS turn-key solutions, or can integrate the Eclipse module into your existing LC system.
Eclipse™ DualTec™ - Tip-injection FFF fluidics control system that integrates with your HPLC pump and autosampler to provide all the advantages of FFF without losing SEC capabilities. Supports switching between any two of AF4, HF5 and SEC.
Eclipse™ AF4™ - Center-downstream injection FFF fluidics control system, optimal when use with HF5 or SEC is not important. The AF4 is required for the fritless channel and semi-preparative FFF.
DAWN® - The most sensitive MALS detector available, anywhere. Incorporates detectors at 18 angles to determine molar masses from 200 Da to 1 GDa and radii from 10 – 500 nm.
- Standard option: ambient temperature
- Heated/cooled option: -15°C to +150°C
- High-temperature option: ambient to +210°C
The DAWN offers special options to handle fluorescent samples: fluorescence-blocking filters and an infrared, 785 nm laser.
miniDAWN® - Second only to the DAWN in sensitivity. Incorporates detectors at 3 angles to determine molar masses from 200 Da to 10 MDa and radii from 10 – 50 nm. Ambient only.
Dynamic Light Scattering Detector
WyattQELS™ - A dynamic light scattering (DLS) module which integrates into a DAWN or miniDAWN MALS detector to provide simultaneous DLS measurements in the same scattering volume.
Refractive Index Detectors
Optilab® - A unique on-line differential refractometer for measuring concentration of any macromolecule, regardless of chromophores. Temperature controlled from 4°C to 65°C.
ECLIPSE - Controls the Eclipse DualTec and AF4 Field-Flow Fractionation (FFF) systems. Coupled with ASTRA for data collection and analysis, ECLIPSE provides users with a turn-key solution for macromolecular characterization.
ASTRA - Our comprehensive software solution for MALS and DLS analysis in chromatography, FFF or batch mode. ASTRA is available in a 21CFR(11) compliant version and offers additional options such as particle analysis and a research database.
"Easy to use"
"We used Wyatt Eclipse AF4 coupled with online MALS and DLS for liposome, nanoparticle, and other formulation characterization. It is easy to use, and is able to produce high quality results." Yingwen Hu, Leidos Biomedical Inc.
"Without this lot quality control checks for the product would have not possible."
"This is the only technology available for absolute molecular size determination with ease of use. It is highly reproducible and recommended for polysaccharide and conjugate vaccine manufacturers." Rajendar Burki, Biological E Limited
"This instrument has proved invaluable for the analysis of very difficult samples."
"This system is able to separate ultra high molecular weight polymers as well as highly branched polymers. Additionally, we are able to determine the radius of different sized nanoparticles and colloids." Stacy Lewis, 3M (Pace Analytical)
"I had good results"
"Asymmetrical flow field-flow fractionation when connected to suitable detector such as multi-angle light scattering (MALS) has shown to be a great technique for the analysis molar mass and root mean square of many food macromolecules. The results are reproducible and with good quality." Catalina Fuentes, Lund University
Interested in other techniques or combining other techniques with FFF?
SEC-MALS: Standard in protein, biopolymer and synthetic polymer characterizations labs around the world, Wyatt MALS detectors are valued for reliable and robust measurements.
CG-MALS: Apply CG-MALS to characterize self- and hetero-association, binding affinity from pM to mM, absolute molecular stoichiometry (not just mole ratios) and more.
DLS: Measure the translational diffusion coefficients Dt of nanoparticles and colloids in solution by quantifying dynamic fluctuations in scattered light. DLS is suitable for ensemble measurements ranging from Rh values of 0.2 nm up to 5,000 nm.
MP-PALS: Wyatt Technology's breakthrough Massively Parallel Phase Analysis Light Scattering (MP-PALS) extends robust ELS measurements to proteins and other biomolecules in native buffer solutions. MP-PALS does everything conventional zeta potential instruments can do and much more.
SEC-IV: SEC-IV may be the optimal means of characterizing materials not amenable to SEC-MALS analysis. By simply adding a DAWN, transform a basic SEC-IV setup into a powerful SEC-MALS-IV polymer characterization station, to analyze absolute determination of molar mass and size regardless of conformation, branched polymers, copolymers and measure Mark-Houwink coefficients ab initio.