Bead conjugation buffer

Manufactured by Quanterix

Sourced in United States

Bead Conjugation Buffer is a laboratory reagent used to facilitate the conjugation of biomolecules, such as proteins or antibodies, to magnetic beads or other solid supports. The buffer provides an optimized chemical environment to promote stable and efficient covalent or non-covalent coupling of the target molecules to the bead surface.

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Lab products found in correlation

1 ethyl 3 3 dimethylaminopropyl carbodiimide hydrochloride edc, by Thermo Fisher Scientific (2 mentions) Z1 particle counter, by Beckman Coulter (1 mentions) Bead wash buffer, by Quanterix (1 mentions) Bead diluent, by Quanterix (1 mentions) Cd63 ab59479, by Abcam (1 mentions)

3 protocols using bead conjugation buffer

Paramagnetic Bead Conjugation Protocol

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For conjugation, 4.2 × 10⁸ paramagnetic carboxylated
beads (Homebrew Singleplex beads, Quanterix) were washed three times
with 300 μL of Bead Wash Buffer (Quanterix) and two times with
300 μL of cold Bead Conjugation Buffer (Quanterix) before resuspending
in 291 μL of cold Bead Conjugation Buffer. The carboxyl groups
on the beads were activated by adding 9 μL of freshly dissolved
1-ethyl-3-(3-(dimethylamino)propyl) carbodiimide hydrochloride (EDC)
and shaken at 4 °C for 30 min. The beads were then washed once
with 300 μL of cold Bead Conjugation Buffer and resuspended
in 300 μL of 0.167 mg/mL capture antibody (MAB62741, R&D
Systems) in cold Bead Conjugation Buffer. Antibody conjugation was
carried out by shaking the beads at 4 °C for 2 h. The beads were
then washed twice with 300 μL of Bead Wash Buffer before resuspending
in 300 μL of Bead Blocking Buffer (Quanterix) and shaking at
room temperature for 30 min. After blocking, the beads were washed
with 300 μL of Bead Wash Buffer and 300 μL of Bead Diluent
(Quanterix) and resuspended in Bead Diluent for storage at 4 °C.
The beads were counted with a Beckman Coulter Z1 Particle Counter.

Kang X., Wu C., Alibakhshi M.A., Liu X., Yu L., Walt D.R, & Wanunu M. (2023). Nanopore-Based Fingerprint Immunoassay Based on Rolling Circle Amplification and DNA Fragmentation. ACS Nano, 17(6), 5412-5420.

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Antibody-Based EV Detection Assays

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Antibodies against EV biomarkers CD63, CD9 and Epcam were selected and prepared for capture and detectionfollowingthe manufacturer’s recommendations. By using commercialEV standards from the HCT116 cell line (HBM-HCT-30/5, Hansabiomed), different antibody combinations were compared and those with the best signals were selected. The antibodies used in the assayswere CD9 MA1-19,002 (ThermoFisher), CD63 ab59479 (Abcam) and Epcam MAB9601 (R&D Systems).
The preparation of SiMoa homebrew kits for EV detection followed the manufacturer’s guideline. In brief, the capture antibody concentration was adjusted to 0.2 mg/mL with Bead Conjugation Buffer (Quanterix) andparamagnetic carboxylated microparticles (Quanterix) were activated with 0.3 mg/mL 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) (Thermo Fisher Scientific, Waltham, MA, USA). To start the biotinylation reaction, 3 μL of the biotin solution (2 mg NHS-PEG4-Biotin dissolved in 383 μL ddH₂O) were added to 100 μL of the detection antibody solution (1.0 mg/mL). The concentration of the recovered antibody was adjusted to 0.2 mg/mL and beads were stored at 4°C.
Finally, two different EVs detection assays were developed: the first assay detecting universal EVs via CD9-CD63 and the second assay detecting tumour-derived EVs via Epcam-CD63.

Wei P., Wu F., Kang B., Sun X., Heskia F., Pachot A., Liang J, & Li D. (2020). Plasma extracellular vesicles detected by Single Molecule array technology as a liquid biopsy for colorectal cancer. Journal of Extracellular Vesicles, 9(1), 1809765.

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Simoa SCC-Ag Sandwich Immunoassay Development

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Capture antibody (rabbit anti-human SerpinB3, 13,218-RP01) and detection antibody (rabbit anti-human SerpinB3, 13,218-T52) for the development of the Simoa SCC-Ag sandwich immunoassay were purchased from Sino Biological (Beijing, China). The preparation of beads with capture and detection antibodies followed the manufacturer’s protocol (Quanterix). The capture antibody concentration was adjusted to 0.2 mg/mL with Bead Conjugation Buffer (Quanterix), and then paramagnetic carboxylated microparticles (Quanterix) were activated with 0.3 mg/mL 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) (Thermo Fisher Scientific, Waltham, MA, USA). To start the biotinylation reaction, 3 μL of the biotin solution (2 mg of NHS-PEG4-Biotin dissolved in 383 μL of ddH₂O) was added to 100 μL of the detection antibody solution (1.0 mg/mL). The concentration of the recovered antibody was adjusted to 0.2 mg/mL, and beads were stored at 4 °C.

Ye S., Sun X., Kang B., Wu F., Zheng Z., Xiang L., Lesénéchal M., Heskia F., Liang J, & Yang H. (2020). The kinetic profile and clinical implication of SCC-Ag in squamous cervical cancer patients undergoing radical hysterectomy using the Simoa assay: a prospective observational study. BMC Cancer, 20, 138.

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