Antibody Technology Grants and Publications

Antibody Technology Shared Resource Grant Descriptions

Short Grant Description

Antibody discovery is based upon highly optimized hybridoma technology performed at a large scale.   Advanced screening techniques include conducting the primary screen by our IQ screener plus high throughput flow cytometer using both cells and protein attached to beads.  In this way we can screen against as many as 20 different proteins and 6 different cell lines simultaneously.  Robotics in the form of our Tecan EVO and Agilent Bravo liquid handlers in combination with the highly multiplexable screening allows us to operate very efficiently at large scales.  An alternate workflow in which cells are seeded into semi-solid media and grown into colonies picked out by our Clone Pix II robot allow us to tackle discovery projects that require clonality prior to screening and allow for efficient subcloning.  Efficient antibody purification and size exclusion chromatography is achieved by use of our AKTA Pure HPLC.  High throughput kinetics and binning is performed by our Carterra high throughput SPR instrument.  We also sequence antibody variable regions using the advanced RACE method.  In addition to the services we provide, we are pleased to provide antibody related expertise from our knowledgeable and experienced staff with respect to any antibody related question.

Long Grant Description 

The Antibody Technology shared resource is staffed by a multidisciplinary team of cell biologists and molecular biologists with over 50 years of combined academic and industrial research experience. The team consists of the shared resource director and three scientific staff who work closely with several academic and industrial clients on a broad range of novel antibody discovery campaigns. Over the past eight years, the core has designed and operated a state-of-the-art antibody discovery platform and gained international recognition as a leader in the field.   The team runs approximately 40 discovery campaigns annually with a successful portfolio of novel antibody compositions for therapeutic and diagnostic applications. The lab occupies 2000 square feet in the Fred Hutch Eastlake Building and has been recently updated with over $2.5 million in new instrumentation. (Instrumentation: Tecan EVO, AATI Infinity, Molecular Devices ClonePix 2, IntelliCyt iQue Screener PLUS, Agilent Bravo, ÄKTA Pure, ProteinSimple WES, Carterra LSA).  Services we offer fall into 4 categories.

Antibody discovery - This activity centers around discovering novel antibodies to a target.  First, we consult with the researcher concerning what characteristics the researcher would like for their antibody to have and how to tailor the immunization, screening and overall project to maximize the chances of being able to achieve an antibody with those characteristics.  This is followed by immunization which we do with KLH conjugated peptides, proteins or cells expressing particular antigens in mice, rats, guinea pig or rabbit.    Advanced screening techniques include conducting the primary screen by our IQ screener plus high throughput flow cytometer using both cells and protein attached to beads.  In this way we have screen against as many as 20 different proteins and 6 different cell lines simultaneously.  Robotics in the form of our Tecan EVO and Agilent Bravo liquid handlers in combination with the highly multiplexable screening allows us to operate very efficiently at large scales.  An alternate workflow in which cells are seeded into semi-solid media and grown into colonies picked out by our ClonePix II robot allow us to tackle discovery projects that require clonality prior to screening.  ClonePix II also allows for efficient subcloning of hybridoma.

Antibody production – Antibody production starts with weening hybridoma for growth in serum free media (SFM). We then expand the hybridoma to volumes ranging from .5L to 20L volumes in SFM.  We purify on ether protein A or Protein G columns using an AKTA Pure HPLC.  We generate anything from mg to gram quantities with an average of 24mg/L of antibody.  We also generate antibody from transiently transfected 293F cells and any stable cell line desired.  In addition, we test the purity of the antibody we generate by size exclusion chromatography.

Antibody characterization – Using our Carterra high throughput SPR platform we generate Kinetic data on up to 384 antibodies in a single run.  Again, using the Carterra we can perform binning on up to 384 antibodies which determines which antibodies block the binding of which antibodies.  The effect of binning is to group the antibodies according to the region which they bind the antigen.  We also have multiple different methods of antibody sequencing including standard cDNA sequencing but also RACE sequencing that use antibody specific primers for reverse transcription.

Advice and training – A large part of what we do is to provide the researchers advice based on our significant expertise at every stage of their antibody discovery project.  This would include target selection, method of immunization and screening.  We also provide advice on what screens they need to perform as well as how to develop assays.  We also help interpret results and provide insight into screening assays.