A Liquid-Handling Robot for Automated Attachment of Biomolecules to Microbeads

Diagnostics, drug delivery, and other biomedical industries rely on cross-linking ligands to microbead surfaces. Microbead functionalization requires multiple steps of liquid exchange, incubation, and mixing, which are laborious and time intensive. Although automated systems exist, they are expensive and cumbersome, limiting their routine use in biomedical laboratories. We present a small, bench-top robotic system that automates microparticle functionalization and streamlines sample preparation. The robot uses a programmable microcontroller to regulate liquid exchange, incubation, and mixing functions. Filters with a pore diameter smaller than the minimum bead diameter are used to prevent bead loss during liquid exchange. The robot uses three liquid reagents and processes up to 107 microbeads per batch. The effectiveness of microbead functionalization was compared with a manual covalent coupling process and evaluated via flow cytometry and fluorescent imaging. The mean percentages of successfully functionalized beads were 91% and 92% for the robot and manual methods, respectively, with less than 5% bead loss. Although the two methods share similar qualities, the automated approach required approximately 10 min of active labor, compared with 3 h for the manual approach. These results suggest that a low-cost, automated microbead functionalization system can streamline sample preparation with minimal operator intervention.

[1]  T. Sulchek,et al.  Bifunctional Janus microparticles with spatially segregated proteins. , 2012, Langmuir : the ACS journal of surfaces and colloids.

[2]  John E. Snawder,et al.  Comparison of a Multiplexed Fluorescent Covalent Microsphere Immunoassay and an Enzyme-Linked Immunosorbent Assay for Measurement of Human Immunoglobulin G Antibodies to Anthrax Toxins , 2004, Clinical Diagnostic Laboratory Immunology.

[3]  T. Sulchek,et al.  Decoupling Internalization, Acidification and Phagosomal-Endosomal/lysosomal Fusion during Phagocytosis of InlA Coated Beads in Epithelial Cells , 2009, PloS one.

[4]  M. Beach,et al.  Multiplex bead assay for serum samples from children in Haiti enrolled in a drug study for the treatment of lymphatic filariasis. , 2011, The American journal of tropical medicine and hygiene.

[5]  J. Guarner,et al.  An in vitro model of the leukocyte interactions associated with granuloma formation in Mycobacterium tuberculosis infection , 2007, Immunology and cell biology.

[6]  S. Nie,et al.  Molecular profiling of single cancer cells and clinical tissue specimens with semiconductor quantum dots , 2006, International journal of nanomedicine.

[7]  D. Selkoe,et al.  Bartels & Selkoe reply , 2013, Nature.

[8]  Jamie M. Heinzman,et al.  Robotic Liquid Handlers and Semiautomated Cell Quantification Systems Increase Consistency and Reproducibility in High-Throughput, Cell-Based Assay , 2010 .

[9]  Mauro Ferrari,et al.  Nanomedicine in cancer therapy: Innovative trends and prospects , 2011, Cancer science.

[10]  William L. Bigbee,et al.  Multiplexed Immunobead-Based Cytokine Profiling for Early Detection of Ovarian Cancer , 2005, Cancer Epidemiology Biomarkers & Prevention.

[11]  T. Sulchek,et al.  TUNABLE COMPLEMENT ACTIVATION BY PARTICLES WITH VARIABLE SIZE AND Fc DENSITY. , 2013, Nano LIFE.

[12]  M. Quinn,et al.  Isolation and Characterization of Tumor Cells from the Ascites of Ovarian Cancer Patients: Molecular Phenotype of Chemoresistant Ovarian Tumors , 2012, PloS one.

[13]  M. Ferrari,et al.  Molecular‐targeted nanotherapies in cancer: Enabling treatment specificity , 2011, Molecular oncology.

[14]  Yuan Qi,et al.  Cell Line Derived Multi-Gene Predictor of Pathologic Response to Neoadjuvant Chemotherapy in Breast Cancer: A Validation Study on US Oncology 02-103 Clinical Trial , 2012, BMC Medical Genomics.

[15]  Philip N Duncan,et al.  Pneumatic oscillator circuits for timing and control of integrated microfluidics , 2013, Proceedings of the National Academy of Sciences.

[16]  Philip N Duncan,et al.  Semi-autonomous liquid handling via on-chip pneumatic digital logic. , 2012, Lab on a chip.

[17]  J. T. Crawford,et al.  Transfer of a Mycobacterium tuberculosis Genotyping Method, Spoligotyping, from a Reverse Line-Blot Hybridization, Membrane-Based Assay to the Luminex Multianalyte Profiling System , 2004, Journal of Clinical Microbiology.

[18]  E. Taioli,et al.  Endometrial cancer: multiplexed Luminex approaches for early detection. , 2008, Expert opinion on medical diagnostics.

[19]  Jörg Maser,et al.  Nanoparticles for Applications in Cellular Imaging , 2007, Nanoscale research letters.

[20]  Bo Chen,et al.  Janus particles as artificial antigen-presenting cells for T cell activation. , 2014, ACS applied materials & interfaces.

[21]  T. Sulchek,et al.  Effects of Microparticle Size and Fc Density on Macrophage Phagocytosis , 2013, PloS one.

[22]  G. Øye,et al.  Plasmachemical Amine Functionalization of Porous Polystyrene Beads: The Importance of Pore Architecture , 2002 .

[23]  E. Dickerson,et al.  Selective removal of ovarian cancer cells from human ascites fluid using magnetic nanoparticles. , 2010, Nanomedicine : nanotechnology, biology, and medicine.