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ADC Conjugation Kit — Conjugation with Speed, Precision, and Reliability

Antibody-Drug Conjugates (ADCs) are targeted therapeutics structured as "Antibody-Linker-Payload". By precisely conjugating cytotoxic payloads (e.g., MMAE, DXd) to antibodies, ADCs enable targeted delivery of toxins to tumor cells. Conjugation technology critically influences ADC efficacy, safety, and manufacturability.
We have a new ADC Conjugation Kit, delivering a rapid, precise, and robust ADC preparation process. Generate functional ADCs within 3 hours while maintaining antibody activity and high recovery. The resulting conjugates are suitable for direct use in cellular or animal studies to accelerate candidate ADC screening.
Technology & Principle
Based on interchain cysteine conjugation technology, this kit reduces antibody disulfide bonds to generate reactive cysteine residues for non-site-specific maleimide conjugation. This classic strategy enables rapid, controlled ADC preparation—requiring only 3h to obtain ADC conjugates with homogeneous DAR. The resulting conjugates demonstrate significant cytotoxicity, offering a robust tool for ADC screening and mechanistic studies.
Technology & Principle
Product Features

Fast: Complete ADC prep in <3 hours

Precise: Controlled DAR 4/8 with preserved antibody activity

Efficient: >95% conjugation, >80% recovery

Low Input: Compatible with ≥2 mg/mL antibodies

Simple: 15 min purification via spin column

Protocol Overview

1. Prepare antibody solution for 2 mg/mL in 1×PBS buffer (pH7.2-7.4)

2. Antibody reduction 37°C, 1h

3. Antibody conjugation 23°C, 1h

4. Reaction quenching RT, 0.5h

5. Purification and buffer change

Product List

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Components

The ADC Conjugation Kit (MMAE, DAR4, 200μg, for human IgG1) (Catalog No.: ADC-P013) as an example.
Cat.No. Components ID Components Size (1 Kit) Storage
Box A
ADC-P013 		C01 Reducing reagent 1 vial -20℃
C02 VcMMAE 1 vial -20℃
C03 Quencher reagent 1 vial -20℃
Box B
ADC-P013-1 		Buffer01 1xPBS (pH7.2-7.4), 11%Trehalose 5 mL 4℃
Desalting column Desalting Spin column, 0.5mL 1 per 4℃

Validation Data

The following ADC products were prepared using the ADC Conjugation Kit (MMAE, DAR4, 200μg, for human IgG1, Catalog No.: ADC-P013).
Antibody resource: Trastuzumab biosimilar

DAR and Purity Validation by HPLC (HIC + SEC)

DAR and Purity Validation by HPLC (HIC + SEC)
DAR and Purity Validation by HPLC (HIC + SEC)

Figure 1. The ADC was prepared using the ADC Conjugation Kit (MMAE, DAR4) and analyzed by HIC and SEC-HPLC. The average drug-antibody ratio (DAR) is 4.0±0.5, and the purity of the ADC is greater than 95%.

DAR Validation by Mass Spectrometry

DAR Validation by Mass Spectrometry

Figure 2. The DAR (3.82) was calculated from the weighted average of the deconvoluted MS peak areas using LC-MS/MS. The results showed the deconvoluted mass spectra of light chains and heavy chains, and the increase in molecular weight caused by the conjugated payload (1316±3 Da). The heterogeneity in N-glycosylation of heavy chain adds to the complexity of the mass spectrum.

Validation of Antigen Binding Activity

Validation of Antigen Binding Activity

Figure 3. Binding affinity of anti-Her2 antibody and anti-Her2 antibody–MMAE conjugate to human Her2 (Cat. No. HE2-H5225) as determined by BLI (Bio-Layer Interferometry). The conjugate exhibits nanomolar affinity (0.57 nM) for human Her2, comparable to the naked antibody (0.86 nM).

In Vitro Cytotoxicity Validation

In Vitro Cytotoxicity Validation

Figure 4. In vitro cytotoxicity assays: The ADC can bind and internalize in target cells (SK-BR-3) with high expression of Her2 and release MMAE inside the cells to induce a cytotoxic effect (IC50=0.0058 µg/mL). Meanwhile, no cytotoxicity was observed in Her2 receptor-negative cell lines (MDA-MB-231).

Performance Consistency Across Kit Sizes

In Vitro Cytotoxicity Validation

Figure 5. In vitro cytotoxicity of products prepared by different size of the ADC Conjugation Kit (Cat. No. ADC-P013, ADC-P014). The result shows very high consistency (RSD＜10%).

Stability Validation

In Vitro Cytotoxicity Validation

Figure 6. The MMAE-ADC was prepared using the ADC Conjugation Kit, which were left at 37℃ for 21 days. The ADCs were analyzed by HIC and SEC-HPLC. The average drug-antibody ratio (DAR) is 4.0±0.5, and the purity of the ADC is greater than 95%.

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Resource Sharing

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Addressing Challenges in Antibody-Drug Conjugate Development

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Building the Perfect Antibody-based Therapeutic from Selection to Engineering and Manufacturing

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FAQ

Q

What is non-site-specific ADC conjugation based on interchain cysteine reduction?

Non-site-specific ADC conjugation using interchain cysteine reduction involves reducing antibody interchain disulfide bonds to generate reactive cysteine thiols, followed by conjugation with maleimide-activated payload-linkers. Unlike site-specific methods that rely on predefined conjugation sites, this approach utilizes naturally occurring reactive residues on antibodies. The resulting ADCs contain a distribution of drug-to-antibody ratio (DAR) species that can be characterized using analytical methods such as HIC-HPLC or LC-MS.
Q

How do MMAE, MMAF, and Exatecan differ as ADC payloads?

MMAE and MMAF are auristatin-class payloads that interfere with microtubule function, while Exatecan is a camptothecin-derived topoisomerase I inhibitor with a distinct mechanism of action. These payloads represent different classes of ADC cytotoxic agents and can be conjugated to antibodies for ADC research applications. ACROBiosystems provides non-site-specific ADC conjugation kits for MMAE, MMAF, and Exatecan payloads.
Q

What DAR does each ADC conjugation kit deliver?

The MMAE and MMAF ADC conjugation kits are designed to generate ADCs with a target DAR of 4, while the Exatecan ADC conjugation kit is designed for a target DAR of 8. After conjugation, DAR can be evaluated using analytical methods such as HIC-HPLC, and ADC purity can be assessed using methods such as SEC-HPLC. These characterization results help evaluate conjugation consistency and ADC quality attributes.
Q

What is the non-site-specific ADC conjugation workflow and how long does it take?

The ADC conjugation workflow includes antibody reduction, payload-linker conjugation, quenching, and purification steps. Using the ADC Conjugation Kit, the complete workflow can be completed in approximately 3 hours, with conjugation efficiency and antibody recovery evaluated after preparation. The streamlined workflow enables preparation of ADC conjugates for downstream characterization and functional studies.
Q

What types of experiments can non-site-specific ADC conjugation kits support?

Non-site-specific ADC conjugation kits can support ADC candidate preparation for early-stage research, including antibody–payload combination evaluation, cytotoxicity studies, binding activity analysis, and preliminary stability assessment. ACROBiosystems provides ADC Conjugation Kits with MMAE, MMAF, and Exatecan payload options, supporting ADC preparation for different antibody and payload combinations.
Q

What reagents are needed for ADC characterization after conjugation?

ADC characterization after conjugation commonly involves evaluating target binding, internalization, and payload-related properties. ACROBiosystems provides complementary reagents including ADC target proteins, internalization detection reagents, linker-cleaving proteases, and anti-payload antibodies to support downstream ADC characterization workflows.
Q

How consistent are the ADCs produced across different kit sizes, and can they be directly used in cell killing assays?

The kit is engineered for high operational robustness. Comparative validation data shows exceptional batch-to-batch and scale-to-scale consistency between the 200 μg (e.g., Cat. No. ADC-P013) and 1 mg (e.g., Cat. No. ADC-P014) kit formats, with in vitro cell cytotoxicity variations exhibiting a relative standard deviation (RSD<10%). Because unreacted free toxin is thoroughly removed during column purification, the resulting ADCs can be added directly to cell-based killing assays, target internalization assays, or animal models without background payload toxicity.
Q

What critical biophysical quality attributes make ACROBiosystems' anti-idiotypic antibodies highly reliable across multi-year clinical campaigns?

Multi-year clinical trials require reagents with unwavering batch consistency and minimal risk of assay drift. ACROBiosystems’ anti-idiotypic antibody portfolio features: Picomolar Affinity (KD): Characterized via Surface Plasmon Resonance (SPR), reaching picomolar binding strength (e.g., clone ADB-Y19 binds Adalimumab with an affinity constant of 1.36 pM). SEC-MALS Verification: Select clones undergo Size-Exclusion Chromatography coupled with Multi-Angle Light Scattering (SEC-MALS) to confirm purity and a defined monomeric molecular weight. High Thermal & Freeze-Thaw Stability: Validated through accelerated 37°C stress tests and repeated freeze-thaw cycles without activity loss.
  • Background
  • Product List
  • Components
  • Validation Data
  • Related Recommendations
  • Resource Sharing
  • Promotion
  • FAQ