oYo-Link® DBCO

For the site-specific labeling of DBCO to the heavy chain of compatible antibodies with up to two DBCO labels per antibody.

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oYo-Link® DBCO Click Chemistry Reagents

Research use only. 

oYo-Link® DBCO enables site-specific labeling to the heavy chain of compatible antibodies with up to two DBCO labels per antibody.  oYo-Link® enables the click reaction to take place prior to antibody labeling which results in highly efficient click reactions. Simply mix oYo-Link DBCO with desired antibody followed by photocrosslinking for conjugation in 30-sec hands on time, 2 hours total.

Requirements: Photo-Crosslinking Device

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NOTE: It is important to perform the click-reaction with Azide-labeled molecule of choice first, prior to antibody conjugation. UV illumination will damage the DBCO activity and thus antibody conjugation should only be performed after the click-reaction.

oYo-Link® DBCO allows for the site-specific labeling of oYo-Link®-compatible antibodies with up to two dibenzocyclooctyne (DBCO) labels per antibody.

oYo-Link enables attachment of the DBCO labels to the heavy chains of the antibody and therefore ensures that the DBCO label does not interfere with antibody binding. Furthermore, site-specific labeling limits 1-2 DBCO labels to be attached per antibody, providing precise antibody-conjugate ratios.

With oYo-Link®, the click reaction takes place prior to antibody labeling which results in highly efficient click reactions.

Only 30 seconds hands-on time, 2 hours total.

The antibody labeling procedure simply requires mixing oYo-Link® -DBCO with the desired antibody followed by photocrosslinking with non-damaging black light (365 nm). Photocrosslinking can be carried in most buffers, including those containing amine-containing molecules (e.g. TRIS, glycine) or storage proteins (see Buffer Compatibility Table).

Applications: Copper-free Click Chemistry.

Requirements: To label your compatible antibodies with oYo-Link®, you will need a light source emitting at 365nm (6-10W). Purchase AlphaThera’sLED Photo-Crosslinking Device here, or use a compatible device.

Placing an order: For credit card purchases, proceed with checkout on website. To request a quote click here.

Technical Support: For any technical questions please contact our technical support team here.

 

Webinar: Simple, fast, site-specific antibody labeling with oYo Link®

Watch our short introduction to AlphaThera’s oYo-Link® site-specific antibody conjugation technology and learn how to streamline your assays with these superior labeling products.

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Antibody Conjugation User Manual

Read our antibody conjugation usual manual for instructions on how to easily conjugate your antibody with oYo-Link products.

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Click Chemistry Protocol with oYo-Link DBCO

Guides users on how to perform copper-free click-chemistry coupling reactions with oYo-Link DBCO.

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Demo: oYo-Link® Antibody Conjugation

Find out how to site-specifically label your antibody with just two steps: mix and illuminate. 30 seconds hands-on time, 2 hours total.

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oYo-Link® Product Brochure

Download our product brochure

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oYo-Link® DBCO Antibody Crosslinking Data

oYo-Link DBCO Photo-crosslinking Efficiency shown by SDS-PAGE: using a 4-12% Tris Gel shows an upwards shift of the antibody heavy chain following site-specific photo-crosslinking with oYo-Link. *Note the size and migration for each unreacted oYo-Link product will differ.

Note: To label mouse IgG1 antibody, you must order the oYo-Link mIgG1 products. This will only label mIgG1 antibody.

Catalog Amount Labeled Price(USD) Spec Sheet
AT3003-100 Labels 100 µg Antibody $300 Download Here
AT3003-mIgG1-100 Label 100 µg mouse IgG1 Antibody $300 Download Here
AT3003-500 Label 500 µg Antibody $1050 Download Here
AT3003-mIgG1-500 Label 500 µg mouse IgG1 Antibody $1050 Download Here
AT3003-1000 Label 1000 µg Antibody $1500 Download Here
AT3003-mIgG1-1000 Label 1000 µg mouse IgG1 Antibody $1500 Download Here

Specifications:

Product Name

oYo-Link® DBCO
Catalog #

AT3003
Lot #

See Product Spec Sheet shipped with product.
Antibody and Buffer Compatibility

View the oYo-Link Antibody & Buffer Compatibility Table here.

Note: To label mouse IgG1 antibody, you must order the oYo-Link mIgG1 products, which will only label mIgG1 antibody.
Molecular Weight

8000 g/mol
Concentration

270 µg/mL
Molar Concentration

33 µM
Storage Buffer

1 x PBS
Shipping Condition

This product is shipped as a dried clear or white pellet with cold packs.
Reconstitution

Before opening, briefly centrifuge each tube to ensure that the entire pellet is at the bottom. For all product tubes: Add 100 µL sterile ddH2O and vortex tube briefly until fully dissolved.
Storage and Stability

The dried product should be stored at -20°C upon receipt. Following reconstitution, DBCO loses its reactivity over time due to its oxidation and addition of water to the triple bond. The reconstituted product should be stored at 4°C for immediate use (< 1 week) and -20°C to -80°C for longer time-spans. Limit the freeze-thaw cycles to less than 5.
Conjugation Instructions

After reconstitution, each product tube is capable of labeling up to 100 µg of antibody.NOTE: It is important to perform the click-reaction with Azide-x first, prior to antibody conjugation. UV illumination will damage the DBCO activity and thus antibody conjugation should only be performed after the click-reaction. Please find and follow the Antibody Conjugation User Manual here
Other Notes

Disposal of product should be performed in compliance with all applicable federal, state, and local regulations. This product does not meet hazard classification criteria based on evaluations made by our company under the OSHA Hazard Communication Standard, 29 CFR 1910.1200.

Multiplexed In Situ Imaging of Site-Specific m6A Methylation with Proximity Hybridization Followed by Primer Exchange Amplification (m6A-PHPEA)

Song, M., Wang, J., Hou, J., Fu, T,. Feng, Y., Lv, W., Ge, F., Peng, R., Han, D., Tan, W. (2024) Multiplexed In Situ Imaging of Site-Specific m6A Methylation with Proximity Hybridization Followed by Primer Exchange Amplification (m6A-PHPEA). ACS Nano. Vol 18/Issue 40. pubs.acs.org/doi/abs/10.1021/acsnano.4c08407

Will oYo-Link work with my antibody?

Please check the antibody compatibility table. Note: oYo-Link mIgG1 can be used to label mouse IgG1 antibody. Currently, oYo-Link mIgG1 is only available for certain products. Please check the product webpage. oYo-Link mIgG1 only labels mouse IgG1 and is not compatible with other antibodies.

Can oYo-Link be used to label antibodies in the presence of albumin and/or Tris?

Yes, since oYo-Link specifically bind to the heavy chain of IgG, it works in the presence of both albumin and Tris. oYo-Link is compatible with all common buffers. See the full buffer compatibility table here.

If an oYo-Link product has accidentally been left at room temperature for a week, will it still work?

With the exception of oYo-Link DBCO, all oYo-Link products are shipped as white or clear pellets and are very stable. While we recommend cold storage, they will not exhibit any loss of activity if left at room temperature for a week. oYo-Link DBCO must be shipped and stored cold (-20°C) immediately upon arrival, otherwise there will be a loss in activity.

Does AlphaThera offer conjugation service?

Yes. If you are interested in our conjugation service, please contact us at support@alphathera.com. For custom conjugation services, you will need to send us your antibody. The conjugation and purification fee depends on the size and specifications of each order. Purification is not required for most applications.

How many oYo-Link labels will be conjugated to each antibody?

For most subclasses and species of antibody, oYo-Link will result in the conjugation of 2 labels per antibody (maximum), one label per heavy chain. For example, ~95% of human IgG and Rabbit IgG will have two labels per antibody. However, there are a few antibodies such as mouse IgG2b and goat IgG, where the conjugation is slightly less efficient. In this case, 60 to 80% of the antibody will be labeled and have a mixture of 0, 1 and 2 labels per antibody. See the antibody conjugation efficiency here

How do I know if oYo-Link was successfully conjugated to my antibody?

Antibody conjugation can be checked on SDS-PAGE gel. An example is present in the supporting data on each product page.

Does 365nm UV illumination damage the antibody and its binding affinity?

The conjugated and unconjugated antibody binding affinities have been tested via both ELISA and cell binding assays and don’t show any difference.

Do I need to remove free oYo-Link after conjugating to antibody?

For most immunoassay applications, purification is not necessary. In our standard recommended protocol, the molar ratio of oYo-Link-to-antibody is 5:1. These conditions ensure that the antibody conjugation efficiency reaches its maximum; however there will be a slight excess remaining in your antibody mixture. This is usually removed during immunoassay washing steps. However, if your application requires high antibody-conjugate purity, please contact us for more information on this process and for a detailed protocol.

For oYo-Link click-chemistry products, should I perform the click reaction before conjugation?

When working with oYo-Link Azide (Catalog #: AT3002) and oYo-Link DBCO (Catalog #: AT3003), it is required to perform the click chemistry coupling reactions with the molecule of interest first, prior to photo-crosslinking to an antibody. This is to avoid UV illumination damage to the click moiety and to avoid diluting oYo-Link prior to the reaction to maximize the reaction efficiency. For oYo-Link Tetrazine (Catalog #: AT3004) and oYo-Link Thiol (Catalog #: AT3001); it is recommended to perform the coupling reaction before photo-crosslinking. This is to avoid diluting oYo-Link prior to the coupling reaction to maximize the reaction efficiency.

What is the lowest amount of antibody that can be labeled by oYo-Link reagents?

We recommend working with at least 1 µg of antibody; however, it is possible that even lower amounts of antibody can be labeled.

Can antibodies that are highly dilute be labeled by oYo-Link reagents?

Yes, antibodies can be efficiently labeled even if diluted to concentrations as low as 50 µg/mL.

oYo-Link® Click-Chemistry Advantages

  • Perform click-reactions prior to antibody conjugation for optimal reaction conditions and a 10x increase in reaction efficiency.
  • Site-specific labeling to the Fc-region every time guarantees no interference with antibody-binding.
  • Site-specific & covalent labeling results in uniform products and consistent, predictable results.
  • <30 seconds “hands-on” time, 2 hours total. Two simple steps, Mix and Illuminate.
  • Label as little as 1 µg of antibody per reaction
  • Label diluted antibody solutions, as low as 50 µg/mL
  • Compatible with all common buffers, including BSA, Tris & Azide
  • >90% labeling efficiency of most antibodies
  • After resuspension, aliquot and save for future reactions

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