DNA Analysis Using an Electrospray Scanning Mobility Particle Sizer

Abstract

A scanning mobility particle sizer (SMPS) allows size separation of gas phase particles according to their electrophoretic mobilities. The addition of an electrospray source (ES) recently allowed extension of SMPS analysis to the macromolecular range. We demonstrate here the application of ES-SMPS to nucleic acids analysis. Single- and double-stranded DNA molecules ranging from 6.1 kDa (single-stranded DNA 20 nucleotides in length) to 300 kDa (500 base-pair double-stranded DNA) were separated and detected by ES-SMPS at the picomole to femtomole levels. The measured electrophoretic mobility diameters were found to correlate with the analytes' molecular weights, while the peak areas could yield quantitative information. No fragmentation of DNA was observed under the conditions employed. Different apparent densities were observed for single-stranded and double-stranded DNAs, showing a different behavior for each type of biomolecule. The total analysis time was about 3 min/spectrum. Further optimization of ES-SMPS is expected to make it a fast and sensitive technique for biopolymer characterization.

Analytical Approaches for Size and Mass Analysis of Large Protein Assemblies

“…These diameters can be converted to a mass spectrum as well, due to the generally good correlation between mobility size and molecular weight, although as in IMMS, exceptions to this rule may apply for nonglobular structures, such as shell-like viral capsid intermediates (66,112). The GEMMA analyzer has an extended size range and has been successfully used for particles of sizes ranging from 3 nm to 100 nm, which covers a mass range of a few kDa (small proteins) to 100 MDa (whole viruses or even cell organelles and DNA) (113). Applications of the GEMMA analyzer have ranged from analyzing antibody aggregation, macromolecular protein complexes (114), synthetic polymers (115), intact viruses (116), and lipoparticles (117).…”

Electrospray–differential mobility analysis of bionanoparticles

“…when it was used for determining the mobility diameter of single stranded and double stranded DNA from size range 6 kDa to 900 kDa [46]. Polyethylene glycols (PEGs) in the size range of 4k to 700 kDa were used to establish that molecular weight and mobility diameter could be correlated by a one-third power law [47] a correlation that had already been established for proteins three years prior [11].…”

“…Here ρ is the density of the polymer (or protein), MW is the molecular weight, N avg is the Avagadro's number and d DMA is the analyte diameter determined by DMA [46,50]. It should be pointed out that for a known gas phase density of a particle, equation…”

Comparison of various nano-differential mobility analysers (nDMAs) applying globular proteins

“…Combined with charged particle or molecular ion generation from liquid samples with nano-electrospray sources [6,7] (n-ES) and subsequent neutralization of the highly charged species [8][9][10], this method has already been successfully applied in analysing important biochemical analytes, such as viruses and virus fragments [4,11], bacteriophages, DNA [12], proteins and protein complexes [4,13], as well as synthetic materials, such as polyethylene glycol polymers [14], polystyrene [15], PAMAM dendrimers [16] and nanometer sized silica particles [17].…”