Ez link micro sulfo nhs lc biotinylation kit

Manufactured by Thermo Fisher Scientific

Sourced in United States

The EZ-Link Micro Sulfo-NHS-LC biotinylation kit is a laboratory product designed for the biotinylation of proteins and other biomolecules. It contains the necessary reagents to facilitate the covalent attachment of biotin to target molecules, enabling their detection or purification using streptavidin-based methods.

Automatically generated - may contain errors

Lab products found in correlation

Human ace2 protein, by Sino Biological (1 mentions) Marv angola gpdtm, by IBT Bioservices (1 mentions) Alexa fluor 647 labeling kit, by Thermo Fisher Scientific (1 mentions) 96 well plate, by Thermo Fisher Scientific (1 mentions) Sa pe, by BioLegend (1 mentions) Aria 3 fusion cell sorter, by BD (1 mentions) Apc sa, by BioLegend (1 mentions) Sa bv421, by BioLegend (1 mentions) Lsrfortessa flow cytometer, by BD (1 mentions) Pierce bca assay kit, by Thermo Fisher Scientific (1 mentions) Fluorescence biotin quantitation kit, by Thermo Fisher Scientific (1 mentions) Milli q system, by Merck Group (1 mentions) Igm percp cy5.5 g20 127, by BD (1 mentions) Lsrfortessa cell analyzer, by BD (1 mentions) 7 aminoactinomycin d 7 aad, by Thermo Fisher Scientific (1 mentions) Hla dr bv650, by BioLegend (1 mentions) Cd14 bv510 m5e2, by BioLegend (1 mentions) Igg bv786 g18 145, by BD (1 mentions) Cd20bv605 2h7, by BioLegend (1 mentions)

9 protocols using ez link micro sulfo nhs lc biotinylation kit

Recombinant SARS-CoV-2 Spike RBD Protein Production

Cited 2 times

Check if the same lab product or an alternative is used in the 5 most similar protocols

Human ACE2 protein was ordered from Sino Biological (Beijing, China). Recombinant SARS-CoV-2 spike RBD proteins with His-tag from wild-type virus (wt RBD) as well as the delta variant (delta RBD) were produced by ISAR Bioscience (Planegg, Germany) with the wt RBD being taken from the S protein nucleotide sequence of the SARS-CoV-2 Wuhan Hu-1 genome (GenBank accession number MN908947, positions 22517 to 23183), while the delta RBD contained the following mutations: L452R and T478K. Details on the procedures were published before [45 (link), 46 (link)]. Shortly, CHO cells were transfected with plasmid vectors containing the DNA sequences for the RBD proteins with an added His-tag and subsequently grown at 37 °C. The supernatants were centrifuged and filtered and subsequently purified using HisTrap columns. Protein content after elution was determined by OD₂₈₀ measurement and the relevant fractions were dialyzed.
The biotinylation of wt RBD and human ACE2 was done using the EZ-Link Micro Sulfo-NHS-LC biotinylation kit (Thermo Scientific #21935 or #A39257) with 20-fold molar excess according to the standard procedure instruction, followed by removal of excess biotin by dialysis against 1 L PBS for 16 h at 4 °C, using Slide-A-Lyzer™ Dialysis Cassettes, 7 K MWCO, 0.5 mL (Thermo Scientific #66373).

Klüpfel J., Paßreiter S., Rumpf M., Christa C., Holthoff H.P., Ungerer M., Lohse M., Knolle P., Protzer U., Elsner M, & Seidel M. (2022). Automated detection of neutralizing SARS-CoV-2 antibodies in minutes using a competitive chemiluminescence immunoassay. Analytical and Bioanalytical Chemistry, 415(3), 391-404.

+ Open protocol

+ Expand

Antibody Effector Function Assay

Check if the same lab product or an alternative is used in the 5 most similar protocols

Assays for antibody effector functions were adapted from previously established protocols.²⁰ (link) Post–challenge NHP sera were inactivated by gamma-irradiation (4 MRad)¹⁸ (link) and removed from the maximum containment laboratory according to RML SOPs approved by the RML IBC. Recombinant MARV-Angola GPdTM (IBT Bioservices) was tethered to Fluospheres NutrAvidin-Microspheres yellow-green or red (Thermo Fisher Scientific, Waltham, MA) using the EZ-link Micro Sulfo-NHS-LC-Biotinylation kit (Thermo Fisher Scientific).

O'Donnell K.L., Feldmann F., Kaza B., Clancy C.S., Hanley P.W., Fletcher P, & Marzi A. (2023). Rapid protection of nonhuman primates against Marburg virus disease using a single low-dose VSV-based vaccine. eBioMedicine, 89, 104463.

+ Open protocol

+ Expand

Quantitative Serum Myl3 Assay for Rats

Check if the same lab product or an alternative is used in the 5 most similar protocols

Myl3 was quantitated in rat serum on a custom assay by MPI Research (Mattawan, MI). Samples (5μL) were analyzed in duplicate on Gyrolab™ Bioaffy 1000 CDs (Gyros, Uppsala, Sweden) using the Gyrolab xP workstation with a biotinylated (EZ Link™ Micro Sulfo-NHS-LC biotinylation kit, Thermo Scientific) capture antibody (BiosPacific, Emeryville, CA) and a fluorescently labeled (Alexa Fluor®647 labeling kit, Invitrogen) detection antibody (Abcam, Cambridge, UK). Prior to analysis, samples were diluted 1:4 in Rexxip A buffer (Gyros). Concentrations were interpolated from a 7-point standard curve using a recombinant; his-tagged rat specific myl3 expressed in-house (Eli Lilly and Company, Indianapolis, IN) as a calibrator and fit with a 5-parameter logistic curve. Capture antibody was diluted in PBS + 0.01% Tween 20, detection antibody was diluted in Rexxip F buffer (Gyros), and standards were diluted in Rexxip A. Prior to sample analysis, the performance of the assay was characterized using positive controls (assay calibrator spiked into rat serum) at six different concentrations (approximately mid-way between each standard concentration) ran on 3 different CDs on 3 different days to assess precision and accuracy. Myl3 was quantifiable from 0.05 ng/mL to 200 ng/mL with <12.5 %CV and −5 to −12% relative error.

Kim K., Chini N., Fairchild D.G., Engle S.K., Reagan W.J., Summers S.D, & Mirsalis J.C. (2016). Evaluation of Cardiac Toxicity Biomarkers in Rats from Different Laboratories. Toxicologic pathology, 44(8), 1072-1083.

+ Open protocol

+ Expand

Ribosome Interaction Assay with RACK1

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

Ribosome interaction experiments were performed as described previously (32 (link)). Briefly, 96-well plates (Nunc) were coated overnight at 4°C with 50 μg/well of RACK1-downregulated HeLa cell lysate diluted in 50 μl of PBS. The coating solution was removed, and nonspecific sites were blocked for 1 h at room temperature with 5% BSA in PBS. Plates were washed with 200 μl/well PBS–0.05% Tween 20. Previously purified recombinant proteins (MBP, wild-type RACK1-MBP, or R36D K38E mutant RACK1-MBP) were biotinylated by using the EZ-Link Micro Sulfo-NHS-LC biotinylation kit (Thermo Scientific) according to the manufacturer's instructions. The biotinylated recombinant proteins were added at 0.5 μg per well in 50 μl PBS. The recombinant proteins were incubated on the coated total cell extract for 2 h at room temperature. After washing with PBS–0.05% Tween 20, HRP-conjugated streptavidin in PBS–0.05% Tween 20 was added to wells for 30 min at room temperature in a final volume of 50 μl. After washes, 3,3′,5,5′-tetramethylbenzidine (TMB) was used according to the manufacturer's protocol (Sigma-Aldrich) to detect streptavidin peroxidase activity. HCl at 1 N was used as the stop solution. The absorbance at 450 nm was read on an Infinite F200 multiwell plate reader (Tecan).

Gallo S., Ricciardi S., Manfrini N., Pesce E., Oliveto S., Calamita P., Mancino M., Maffioli E., Moro M., Crosti M., Berno V., Bombaci M., Tedeschi G, & Biffo S. (2018). RACK1 Specifically Regulates Translation through Its Binding to Ribosomes. Molecular and Cellular Biology, 38(23), e00230-18.

+ Open protocol

+ Expand

Quantifying SARS-CoV-2 Antibody Responses

Check if the same lab product or an alternative is used in the 5 most similar protocols

The recombinant SARS-CoV-2 prefusion stabilized S protein and RBD were biotinylated using an EZ-Link Micro Sulfo-NHS-LC Biotinylation Kit (Thermo Fisher Scientific) according to the manufacturer’s instructions. Probes were generated by coupling biotinylated proteins to fluorophore-conjugated streptavidin (SA) molecules for detection by flow cytometry (SA-BV421, SA-APC, and SA-PE, BioLegend). Isolated PBMCs were stained by a panel of antibodies listed in Table S3 as well as S protein and RBD probes (S-APC, S-PE, and RBD-BV421, 100 ng each). The samples were analyzed on a BD Aria III Fusion cell sorter (weeks 6 and 30) or a BD LSRFortessa flow cytometer (all other time points). At weeks 6 and 30, memory B cells (CD3⁻ CD11c⁻ CD14⁻ CD16⁻ CD123⁻ HLA-DR⁺ CD20⁺ IgM⁻ IgG⁺) double-positive for S protein binding were single cell sorted into 96-well plates and frozen immediately on dry ice for subsequent B cell receptor (BCR) amplification. Data were analyzed using FlowJo v.10.7.1 (FlowJo).

Lenart K., Hellgren F., Ols S., Yan X., Cagigi A., Cerveira R.A., Winge I., Hanczak J., Mueller S.O., Jasny E., Schwendt K., Rauch S., Petsch B, & Loré K. (2022). A third dose of the unmodified COVID-19 mRNA vaccine CVnCoV enhances quality and quantity of immune responses. Molecular Therapy. Methods & Clinical Development, 27, 309-323.

+ Open protocol

+ Expand

Biotinylation and Sulfonation of Antibodies

Check if the same lab product or an alternative is used in the 5 most similar protocols

For biotinylation
of antibody EP1536Y, 40 μL of a 0.90 mg/mL solution of the antibody
was incubated with 1.61 μL of 11 mM Sulfo-NHS-biotin (50-fold
molar excess) on ice for 2 h according to the manufacturer’s
protocol (EZ-Link Micro Sulfo-NHS-LC-biotinylation kit, Thermo Fisher
Scientific). Excess free biotin was removed using a 0.5 mL of 7 kDa
MWCO Zeba spin desalting column (Thermo Fisher Scientific). 200 μL
of ultrapure water (purified using a Milli-Q system, Millipore, Burlington,
MA) was added as a stacker volume for elution of the biotinylated
EP1536Y antibody. 40 μL of 1.167-mg/mL solution of antibody
MJFR1 was biotinylated using the same biotinylation kit. Biotinylation
levels were evaluated using a fluorescence biotin quantitation kit
(Thermo Fisher Scientific). For sulfonation, 40 μL of a 0.90
mg/mL of EP1536Y solution was incubated with Sulfo-TAG NHS-ester (MSD)
according to the manufacturer’s instructions. The protein concentration
of all antibody conjugates was determined using a Pierce BCA assay
kit (Thermo Fisher Scientific). The antibody solutions were aliquoted
and stored at 4 °C until use.

Dutta S., Hornung S., Taha H.B., Biggs K., Siddique I., Chamoun L.M., Shahpasand-Kroner H., Lantz C., Herrera-Vaquero M., Stefanova N., Loo J.A, & Bitan G. (2023). Development of a Novel Electrochemiluminescence ELISA for Quantification of α-Synuclein Phosphorylated at Ser129 in Biological Samples. ACS Chemical Neuroscience, 14(7), 1238-1248.

+ Open protocol

+ Expand

Biotinylation of Broadly Neutralizing Antibodies

Check if the same lab product or an alternative is used in the 5 most similar protocols

All bNAbs (3BNC117 (CD4bs), 10–1074 (V3-loop), PG16 (V1V2-loop), 35O22 (gp120-gp41 interface), 10E8 (MPER gp41) were biotinylated with EZ-Link Micro Sulfo-NHS-LC-Biotinylation Kit (Thermo Fisher). Briefly, a total of 2.5 μg bNAb mix of equal quantities was biotinylated with a 30-fold molar excess of Sulfo-NHS-LC-Biotin in 500 μL volume for 60 min at room temperature (RT). To remove excess Biotin, biotinylated bNAbs were purified over a Zeba Spin Desalting Column. Biotinylated bNAbs were stored at 4°C until further use.

Otte F., Zhang Y., Spagnuolo J., Thielen A., Däumer M., Wiethe C., Stoeckle M., Kusejko K., Klein F., Metzner K.J, & Klimkait T. (2023). Revealing viral and cellular dynamics of HIV-1 at the single-cell level during early treatment periods. Cell Reports Methods, 3(6), 100485.

+ Open protocol

+ Expand

SARS-CoV-2 Spike Protein Binding Assay

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

To generate the protein probes, the recombinant WA-1 S-2P and RBD proteins were biotinylated using the EZ-Link Micro Sulfo-NHS-LC Biotinylation Kit (Thermo Fisher Scientific, Waltham, MA, USA) according to the manufacturer’s instructions [20 (link)]. Streptavidin-conjugated fluorophores (SA-PE, SA-APC or SA-BV421) and biotinylated proteins were coupled at a 4:1 molar ratio. The cryopreserved PBMCs were thawed and stained with 100 ng of the fluorescent protein probes for 20 min at 4 °C, followed by staining with 7-aminoactinomycin D (7-AAD, Thermo Fisher, Waltham, MA, USA) and a panel of antibodies, IgM PerCP-Cy5.5 (G20-127, BD, Franklin Lakes, NJ, USA), IgD FITC (polyclonal, Southern Biotech, Birmingham, AL, USA), CD3 BV510 (SP34-2, BD), CD14 BV510 (M5E2, BioLegend, San Diego, CA, USA), CD16 BV510 (3G8, BD), CD20 BV605 (2H7, BioLegend, San Diego, CA, USA), HLA-DR BV650 (L243, BioLegend, San Diego, CA, USA) and IgG BV786 (G18-145, BD, Franklin Lakes, NJ, USA), for another 20 min at 4 °C. After staining, the cells were washed with FACS buffer (PBS supplemented with 2% heat-inactivated fetal calf serum) and fixed with 1% formaldehyde solution. The samples were acquired using a BD LSRFortessa cell analyzer (BD, Franklin Lakes, NJ, USA). The data were analyzed using FlowJo software v.10.7.1 (FlowJo, Ashland, OR, USA).

Sanders J.W., Ewing D., Sundaram A.K., Gamble C.S., Blevins M., Liang Z., Sanders L.A., Ornelles D.A., Sun P., Lenart K., Feuerstein H., Loré K., Petrovsky N., Williams M, & Porter K.R. (2024). Immunogenicity and Protective Efficacy of Psoralen-Inactivated SARS-CoV-2 Vaccine in Nonhuman Primates. Vaccines, 12(5), 451.

+ Open protocol

+ Expand

Biotinylation of Lipopolysaccharide

Check if the same lab product or an alternative is used in the 5 most similar protocols

10 mg LPSBV was biotinylated with EZ-Link Micro Sulfo-NHS-LC-Biotinylation Kit (Thermo Scientific), according to the manufacturer’s protocol, using PBS as a solvent. To remove PBS, an exhaustive dialysis against distilled water was performed. The biotinylated LPS_BV (bioLPS_BV) was then collected and lyophilized. For in vitro experiments, lyophilized bioLPS_BV was dissolved in distilled water in concentrations not higher than 1 mg mL⁻¹.

Steimle A., Michaelis L., Di Lorenzo F., Kliem T., Münzner T., Maerz J.K., Schäfer A., Lange A., Parusel R., Gronbach K., Fuchs K., Silipo A., Öz H.H., Pichler B.J., Autenrieth I.B., Molinaro A, & Frick J.S. (2019). Weak Agonistic LPS Restores Intestinal Immune Homeostasis. Molecular Therapy, 27(11), 1974-1991.

+ Open protocol

+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Revolutionizing how scientists
search and build protocols!