Mouse Monoclonal Antibody Development 


ichorbio scientific staff has extensive monoclonal antibody development experience and offers clients the opportunity to collaborate on the development of antibodies against soluble proteins, cell surface molecules, or haptens for use in detection, neutralization or in vivo depletion. Through the use of proprietary development technologies, we have demonstrated success in developing monoclonal antibodies against single amino acid substitutions, splice variants and for neutralization. Host animals include mice or hamsters.

  • Mouse Monoclonal antibody development timeline1
  • Phase I Immunization and Serum Screening (6-10 Weeks) 1
  • Phase II Splenocyte Fusion and Screening of Parental Cultures (4-6 Weeks) 2
  • Phase III Subcloning and Screening (4-6 Weeks) 2
  • Phase IV In vitro Monoclonal Antibody Production (6-8 Weeks) 2
  • Mouse monoclonal antibody development in vitro production3
  • Mouse monoclonal antibody development: Additional services 3
  • Purification 3
  • Clearance 4
  • Physico Chemical QCs : 4
  • Technical QCs : 4
  • Functional QCs : 4
  • Formatting 4


Mouse Monoclonal antibody development timeline

Phase I Immunization and Serum Screening (6-10 Weeks)

To expedite our client’s development projects and enhance the immunization process, up to ten (10) mice or hamsters may be immunized for each antigen. Immune responses are elicited using immunogens such as transfected cell lines, DNA, recombinant proteins, native proteins, peptides or haptens conjugated to carrier molecules. A variety of adjuvants are available for non cellular immunogens. Test bleeds are taken for purposes of monitoring antibody titer and/or neutralization activity, usually by ELISA. Screening data from immunized animals is presented to clients in graphical form to determine the best animal(s) suited for B-cell fusions. 


Phase II Splenocyte Fusion and Screening of Parental Cultures (4-6 Weeks)

A splenocyte fusion is performed on one or more animals selected from phase I. Splenocytes are isolated and fused with one or more myeloma cell lines (P3X63Ag8.653) using polyethylene glycol. Fused cells are plated at specific densities and grown under selection in HAT media. The parental cultures from the fusion are screened by ELISA or an appropriate application based method and stable positive cultures are subcloned and expanded for cryopreservation. 


Phase III Subcloning and Screening (4-6 Weeks)

Cloning I: Strongly positive parental cultures exhibiting the desired functions derived from fusion cultures are selected for first round cloning using the limiting dilution method. 96 well panels are screened and positive clones are selected for round two limiting dilution cloning.

Cloning II: Positive clones from first round cloning will be subjected to a second round of limiting dilution cloning. Stable clones will be expanded into 24 well plates and then into T- flasks for further characterization. At this time, supernatant from each clone may be sent to clients for further testing and characterization.


Phase IV In vitro Monoclonal Antibody Production (6-8 Weeks)

At no additional cost, clients may select one clone for in vitro production of approximately one liter of supernatant. The supernatant is then purified using protein A chromatography and provided to clients for further characterization.


Mouse monoclonal antibody development in vitro production

Our facilities are equipped to produce from mg to 100g mAb.

For requests of 200mg and more, a test study is systematically performed to evaluate the clone productivity.

ichorbio has extensive experience in large scale mammalian cell culture using steam in place (SIP) stirred tank, packed bed or hollow fiber bioreactors. In cases where projects are still in the proof of concept stage, in vitro production can be performed in roller bottles or shake flasks. Bioreactor production is available at small and large scale in our production facility.

Work is performed in dedicated cleanroom suites using standard operating procedures and manufacturing data is recorded in cGMP compliant batch records. Master manufacturing files are archived and bio-analytical lot release testing records are provided to clients.

A key factor to expedite delivery and reduce production costs is the availability of multiple scale upstream production platforms. For Hybridoma cells where production requirements are less than 500 milligrams, roller bottles may be used. 

For milligram quantities of recombinant proteins, shake flasks are often utilized followed by IMAC or other purification methods. 

However, when larger production quantities are required, steam in place (SIP) stirred tank bioreactors or hollow fiber systems are available in dedicated cleanroom suites for each product.

Based on the requested mAb amount and the clone productivity, we select the best production scheme using flasks, roller bottles or bioreactors.

Mycoplasma diagnostics and cell line decontamination can also be performed.

Mouse monoclonal antibody development: Additional services



Affinity chromatography (protein A,G,L), ion exchange, gel filtration chromatography, IgM purification.



Solutions for DNA and endotoxin removal.


Physico Chemical QCs :

Purity, Quantity, Integrity, Sterility,…


Technical QCs :

Method validation (ELISA, FACS, IHC,…)


Functional QCs :

Binding, cytotoxicity, antibody-dependent cell-mediated cytotoxicity (ADCC), CDC, in vivo testing….

Isotyping, sequencing, serum-free production, cell-line adaptation, cell banking, productivity assessment, variable chains sequencing, cloning for recombinant antibody generation.

ELISA tests can be developed.


Conjugation and freeze-drying services.