Large Molecule Bioanalysis

KCAS is an industry leading expert in Large Molecule Bioanalysis to quantify proteins, antibodies and other large molecule modalities in various fluids and tissues from early discovery through clinical phases.

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Innovative large molecule bioanalysis

Our highly experienced scientists use appropriate state-of-the-art technologies to best support all aspects of large molecule bioanalysis which include various ligand binding platforms or Hybrid LC-MS approaches, as well as Flow cytometry and molecular technologies.

Large Molecule Bioanalysis at KCAS Bio refers to the bioanalysis of any modality with a molecular weight of greater than 10K Daltons. These would include some of the more common biomolecules such as:

• Monoclonal Antibodies (Mabs)
• Antibody Drug Conjugates (ADCs)
• Antibody Oligo/RNA Conjugates (AOC/ARC)
• Antibody fragments
• Fusion Proteins
• Mono/Di/Tri - specifics
• Gene Therapies
• Pegylated Biotherapeutics
• Other Large Molecule modalities

Considerations for LBA vs. LC-MS

Ligand Binding Assays

Historically, bioanalytical support for large molecule development has utilized ligand binding (LBA) techniques. LBA has been the platform of choice for pharmacokinetic, pharmacodynamic and anti-drug antibody analyses relying on the quality of reagents to help provide sensitive and selective assays. We offer both plate-based and cell-based approaches to meet your program's needs.

KCAS Bio has extensive experience in developing and validating large molecule PK and ADA methods using various technologies such as MSD, ELISA, Luminex, SMCxPRO, depending on the sensitivity requirements of the requested assay.

Ligand Binding Assays


LC-MS/MS is another critical technology used for the Bioanalysis of proteins or large molecule biotherapeutics. The improvement in mass spectrometric performance coupled with specific sample preparation approaches has resulted in a growing focus on quantitation of large molecules using LC-MS/MS.

Hybrid or traditional LC-MS/MS provides an ideal complement to the more established ligand-binding approaches. At KCAS Bio, we partner with our clients to identify to "best" solution regardless of technology.

Liquid Chromatography Mass Spectrometry

Hybrid LC-MS/MS

Hybrid LC-MS/MS approaches are increasingly being utilized to support complex biotherapeutic modalities and can be a great alternative to ligand binding assays. Combining the principles of ligand binding for target analyte enrichment with LC-MS/MS provides an alternative platform for quantitation of large molecules and in some cases the preferred approach. LC-MS/MS can provide significant advantages to quantitate large molecules and at KCAS Bio, we have seen a number of examples where LC-MS/MS provided a clear path forward when an LBA approach met significant hurdles. KCAS Bio has supported many modalities using Hybrid LCMS including ADCs, Mabs, Fusion proteins, Bi or Tri-specifics and more recently antibody oligo conjugates (AOCs). We increasingly see that hybrid LC-MS/MS is becoming the technology of choice for many ADCs or AOCs.

Traditional protein LC-MS

Traditional protein LC-MS quantitation typically will involve direct protein digestion (from matrix) followed by more traditional SPE or other sample prep cleanup steps at the peptide level. This approach is advantageous when there are not good reagents available for LBA or Hybrid strategies. The addition of an enzymatic digestion to produce a range of characteristic peptides provides surrogates that are more compatible with the mass range of many quantitative MS instruments.

LBA or Hybrid LC-MS - which is better?

It depends! KCAS Bio is a strong advocate of assessing the relative merits of available technologies to derive to the optimal outcome. Our depth of experience in all bioanalytical methodologies all under one roof needed for large molecule Bioanalysis - combined with a plethora of instrumentation - ensures we take the best approach for our clients. As you prepare your project, consider the below.

Hybrid LC-MS


  • High selectivity/specificity
  • Good sensitivity
  • Less reliant on “good” critical reagents, only need 1 for target analyte clean-up
  • Easily multiplexed
  • Able to differentiate small amino acid differences
  • Addition of internal standards to account for sample-to-sample variability


  • Capital costs
  • Special Expertise
  • Method Development time is variable
  • Sequential Analysis – time



  • High throughput
  • Good sensitivity
  • Capital investment minimal depending on platform


  • Need for 2 reagents (pairs)
  • Time needed to generate the reagents
  • Limited selectivity and ability to multiplex
  • Matrix interferences
  • Potential Lack of Translatability between species and fluids

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