Subunit middle-down analysis

Subunit middle-down analysis

Monoclonal antibody (mAb) subunit analysis involves the antibody fragmentation through digestion and reduction of IgG antibody molecules. The cysteine protease produced by S.pyogenes, known as IdeS, is highly specific. IdeS digestion followed by reduction generates three subunits that are readily separated by reversed-phase (RP) UHPLC using Thermo Scientific MAbPac RP columns. Deconvolution of subunit mass spectrometry (MS) spectra is easy with powerful Thermo Scientific BioPharma Finder software.

Popular subunit mass analysis products

Vanquish UHPLC System

Q Exactive BioPharma Platform

BioPharma Finder

Subunit mass analysis workflow

Sub-Unit Mass Analysis Workflow

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Antibody fragment types
Antibody Fragmentation

Fab Preparation Kit

Thermo Scientific Pierce Fab Preparation Kit uses immobilized papain protease to digest human or mouse IgG antibodies to make separate Fab and Fc antibody fragments and subsequently to purify the Fab using Protein A agarose.


Subunit mass analysis categories

High-resolution separations

Use Thermo Scientific MAbPac RP columns for high-performance, reversed-phase HPLC and LC/MS characterization of mAbs, antibody fragments, charge variants, antibody drug conjugates (ADCs), PEGylated proteins, and bispecific proteins.

Chromatography built for biopharma

The innovative architecture of the new Thermo Scientific Vanquish Flex UHPLC system is tailored to the specific and demanding analytical requirements that these essential drugs impose. Vanquish Flex UHPLC has been designed and manufactured to exacting engineering standards to be truly built for biopharma.

Protein reduction

Choose between research grade dithiothreitol (DTT) or Tris-(2-Carboxyethyl)phosphine, (TCEP) for antibody reduction.

State-of-the-art intact protein and subunit analysis
State-of-the-art intact protein and subunit analysis

The BioPharma Option for Thermo Scientific Q Exactive Plus and Q Exactive HF Hybrid Quadrupole-Orbitrap mass spectrometers provides a complete BioPharma characterization solution. With Protein Mode the extreme resolving power of the Orbitrap mass analyzer ensures isotopic resolution of monoclonal antibody subunits and facilitates middle-down sequencing.

State-of-the-art intact protein and subunit analysis
State-of-the-art intact protein and subunit analysis

The sliding window method and Thermo Scientific Xtract deconvolution algorithm within Thermo Scientific BioPharma Finder software enables monoisotopic mass determination of monoclonal antibody subunits.


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Orbitrap for native mass spectrometry providing Omass with a new perspective in drug discovery.


Featured intact protein analysis resource
Intact and Native Characterization webinars

Join experts from Genentech and Thermo Fisher to get great structural insight into your drug product using the Q Exactive BioPharma platform of mass spectrometers.

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