Highly Efficient Binding of Paramagnetic Beads Bioconjugated with 100 000 or More Antibodies to Protein-Coated Surfaces

Mani, Vigneshwaran; Wasalathanthri, Dhanuka P.; Joshi, Amit; Kumar, Challa V.; Rusling, James F.

doi:10.1021/ac3028257

Cited by 47 publications

(48 citation statements)

References 34 publications

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“…The MPs have high surface area to volume ratio here enabling attachment of 300,000 HRP labels and 40,000 antibodies to provide extremely efficient protein capture (Mani et al, 2012) as well as high sensitivity. Magnetic bead position is controlled magnetically in the online capture chamber for rapid binding of proteins, washing, and delivery to the detection chamber.…”

Section: Discussionmentioning

confidence: 99%

On-line protein capture on magnetic beads for ultrasensitive microfluidic immunoassays of cancer biomarkers

Otieno

Krause

Latus

et al. 2014

Biosensors and Bioelectronics

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110

108

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Accurate, sensitive, multiplexed detection of biomarker proteins holds significant promise for personalized cancer diagnostics. Here we describe the incorporation of a novel on-line chamber to capture cancer biomarker proteins on magnetic beads derivatized with 300,000 enzyme labels and 40,000 antibodies into a modular microfluidic immunoarray. Capture and detection chambers are produced from PDMS on machined molds and do not require lithography. Protein analytes are captured from serum or other biological samples in the stirred capture chamber on the beads held in place magnetically. The beads are subsequently washed free of sample components, and wash solutions sent to waste. Removal of the magnet and valve switching sends the magnetic bead-protein bioconjugates into a detection chamber where they are captured on 8 antibody-decorated gold nanoparticle-film sensors and detected amperometrically. Most steps in the immunoassay including protein capture, washing and measurement are incorporated into the device. In simultaneous assays, the microfluidic system gave ultralow detection limits of 5 fg mL−1 for interleukin-6 (IL-6) and 7 fg mL−1 for IL-8 in serum. Accuracy was demonstrated by measuring IL-6 and IL-8 in conditioned media from oral cancer cell lines and showing good correlations with standard ELISAs. The on-line capture chamber facilitates rapid, sensitive, repetitive protein separation and measurement in 30 min in a semi-automated system adaptable to multiplexed protein detection.

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Section: Discussionmentioning

confidence: 99%

On-line protein capture on magnetic beads for ultrasensitive microfluidic immunoassays of cancer biomarkers

Otieno

Krause

Latus

et al. 2014

Biosensors and Bioelectronics

Self Cite

110

108

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“…52 In brief, proteins, (1 mg/mL) were first immobilized on the carboxyl functionalized gold surface using amine coupling chemistry. Carboxyl groups were activated using a 1:2 mixture of freshly prepared 4 mg EDC (3-( N , N -dimethylamino) propyl- N -ethylcarbodiimide) and 11 mg NHSS ( N -Hydroxysulfosuccinimide) by flowing for 10 min at 50 μ L/min.…”

Section: Methodsmentioning

confidence: 99%

“…Then, the proteins in buffer were immobilized at a flow rate of 20 μ L/min for 1500 s. Unreacted sites were blocked using 1 M glycine at pH 8.0 buffer. 52 …”

Section: Methodsmentioning

confidence: 99%

Antibody-like Biorecognition Sites for Proteins from Surface Imprinting on Nanoparticles

Bhakta

Seraji

Suib

et al. 2015

ACS Appl. Mater. Interfaces

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Natural antibodies are used widely for important applications such as biomedical analysis, cancer therapy, and directed drug delivery, but they are expensive and may have limited stability. This study describes synthesis of antibody-like binding sites by molecular imprinting on silica nanoparticles (SiNP) using a combination of four organosilane monomers with amino acid-like side chains providing hydrophobic, hydrophilic, and H-bonding interactions with target proteins. This approach provided artificial antibody (AA) nanoparticles with good selectivity and specificity to binding domains on target proteins in a relatively low-cost synthesis. The AAs were made by polymer grafting onto SiNPs for human serum albumin (HSA) and glucose oxidase (GOx). Binding affinity, selectivity, and specificity was compared to several other proteins using adsorption isotherms and surface plasmon resonance (SPR). The Langmuir–Freundlich adsorption model was used to obtain apparent binding constants (KLF) from binding isotherms of HSA (6.7 × 104) and GOx (4.7 × 104) to their respective AAs. These values were 4–300 fold larger compared to a series of nontemplate proteins. SPR binding studies of AAs with proteins attached to a gold surface confirmed good specificity and revealed faster binding for the target proteins compared to nontarget proteins. Target proteins retained their secondary structures upon binding. Binding capacity of AAHSA for HSA was 5.9 mg HSA/g compared to 1.4 mg/g for previously report imprinted silica beads imprinted with poly(aminophenyl)boronic acid. Also, 90% recovery for HSA spiked into 2% calf serum was found for AAHSA.

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“…This behavior is consistent with previous findings for magnetic beads decorated with multiple antibodies. 18 …”

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confidence: 99%

Albumin removal from human serum using surface nanopockets on silica-coated magnetic nanoparticles

et al. 2017

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Selective removal of albumin from human serum is an essential step prior to proteomic analyses, especially when using mass spectrometry. Here we report stable synthetic nanopockets on magnetic nanoparticle surfaces that bind to human serum albumin (HSA) with high affinity and specificity. The nanopockets are created by templating HSA on 200 nm silica-coated paramagnetic nanoparticles using mixed polymer layers of 4 organo-silane monomers. These monomers have amino acid-like side chains providing hydrophobic, hydrophilic and H-bonding interactions that closely mimic features of binding sites on antibodies. The binding capacity of the material was 21 mg HSA/g, and consistently removed ~88% albumin from human serum in multiple repeated use.

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Highly Efficient Binding of Paramagnetic Beads Bioconjugated with 100 000 or More Antibodies to Protein-Coated Surfaces

Cited by 47 publications

References 34 publications

On-line protein capture on magnetic beads for ultrasensitive microfluidic immunoassays of cancer biomarkers

On-line protein capture on magnetic beads for ultrasensitive microfluidic immunoassays of cancer biomarkers

Antibody-like Biorecognition Sites for Proteins from Surface Imprinting on Nanoparticles

Albumin removal from human serum using surface nanopockets on silica-coated magnetic nanoparticles

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