Polystyrene Microparticles with Convergently Grown Mesoporous Silica Shells as a Promising Tool for Multiplexed Bioanalytical Assays

Abstract

Functional core/shell particles are highly sought after in analytical chemistry, especially in methods suitable for single-particle analysis such as flow cytometry because they allow for facile multiplexed detection of several analytes in a single run. Aiming to develop a powerful bead platform of which the core particle can be doped in a straightforward manner while the shell offers the highest possible sensitivity when functionalized with (bio)­chemical binders, polystyrene particles were coated with different kinds of mesoporous silica shells in a convergent growth approach. Mesoporous shells allow us to obtain distinctly higher surface areas in comparison with conventional nonporous shells. While assessing the potential of narrow- as well as wide-pore silicas such as Mobil composition of matter no. 41 (MCM-41) and Santa Barbara amorphous material no. 15 (SBA-15), especially the synthesis of the latter shells that are much more suitable for biomolecule anchoring was optimized by altering the pH and both, the amount and type of the mediator salt. Our studies showed that the best performing material resulted from a synthesis using neutral conditions and MgSO4 as an ionic mediator. The analytical potential of the particles was investigated in flow cytometric DNA assays after their respective functionalization for individual and multiplexed detection of short oligonucleotide strands. These experiments revealed that a two-step modification of the silica surface with amino silane and succinic anhydride prior to coupling of an amino-terminated capture DNA (c-DNA) strand is superior to coupling carboxylic acid-terminated c-DNA to aminated core/shell particles, yielding limits of detection (LOD) down to 5 pM for a hybridization assay, using labeled complementary single-stranded target DNA (t-DNA) 15mers. The potential of the use of the particles in multiplexed analysis was shown with the aid of dye-doped core particles carrying a respective SBA-15 shell. Characteristic genomic sequences of human papillomaviruses (HPV) were chosen as the t-DNA analytes here, since their high relevance as carcinogens and the high number of different pathogens is a relevant model case. The title particles showed a promising performance and allowed us to unequivocally detect the different high- and low-risk HPV types in a single experimental run.

Generalized Analysis Approach of the Profile Roughness by Electron Microscopy with the Example of Hierarchically Grown Polystyrene–Iron Oxide–Silica Core–Shell–Shell Particles

“…Particle Synthesis-PS Core Synthesis: PS particles were synthesized by dispersion polymerization as previously reported by us. [13] The polymerization was carried out in a hybridization oven. A solution of 105 mg of ACVA in 10 mL methanol was prepared in an argon atmosphere.…”

“…In particular, micrometer‐sized beads are well suited for the quantitative detection of analytes of interest by single particle‐based (bio)analytical methods because of their rapid reaction kinetics and the requirement of small sample volumes, an area of application we became interested in over the past years. [ 10–14 ] A very promising material combination for single particle‐based assays is achieved by merging the benefits of polystyrene (PS) and silica (SiO 2 ) particles in a core–shell particle format. [ 12,15,16 ]…”

“…In particular, micrometer-sized beads are well suited for the quantitative detection of analytes of interest by single particle-based (bio)analytical methods because of their rapid reaction kinetics and the requirement of small sample volumes, an area of application we became interested in over the past years. [10][11][12][13][14] A very promising material combination for single particle-based assays is achieved by merging the benefits of polystyrene (PS) and silica (SiO 2 ) particles in a core-shell particle format. [12,15,16] Besides equipping CS particles with optical or electrochemical features and grafting (bio)chemical units to their outer surface that drive many of the developments in the field, two less explicit aspects gained increasing attention recently, surface morphology and its impact on functional performance [10,17,18] as well as magnetic properties, strongly facilitating handling and processibility [19,20] especially in view of automated workflows.…”

Analysis of the profile roughness of core-shell microparticles by electron microscopy

“…The particles used in this study were prepared in the frame of the Focus Area project of BAM "MamaLoCA -Modular, multiplexed, antibody-based lab-on-chip analyzer for food control [7]". The following samples have been investigated: i) PS particles synthesized by dispersion polymerization [1], ii) PS particles covered with an iron oxide (Fe 3 O 4 ) layer and iii) CS particles with a PS core covered with a Fe 3 O 4 layer and closed with a second silica (SiO 2 ) shell. The particles have been suspended in ethanol and samples for analysis have been prepared by drop-casting on conventional carbon TEM grids.…”

“…In particular, few micron-sized beads are well suited for the quantitative detection of analytes of interest with their rapid reaction kinetics and requirement of small sample volumes. A very promising material combination for bead-based immunoassays is achieved by merging the benefits of polystyrene (PS) and silica (SiO 2 ) particles in a core-shell (CS) particle format [1,2]. Besides the particle synthesis procedure, characterization is also essential to establish reproducibility and the performance of the particles [3].…”