Studies in veterinary species continue to suffer from the lack of reliable reagents which is exemplified by the paucity of monoclonal antibodies (mAbs) for immunoglobulin (Ig) isotypes and subisotypes. This problem is compounded by the increasing number of Ig being identified as shown by the recent work on the porcine IgG subclasses (click on PUBLICATIONS OF INTEREST). Unfortunately reagents marketed as “anti-porcine IgG1 “ and “anti-porcine IgG2” without supporting evidence, have little meaning in a species with six expressed subclasses. In an effort to begin to resolve problems of this nature, several initiatives are underway.
First, mAbs with well-established specificity are now available through the USDA Center for Veterinary Biologics (CVB) for the cost of shipment. Currently, anti-porcine IgA and anti-IgM are available through this pathway. To read more about this program and to request these mAbs [click on REAGENTS]. These hybridomas were contributed by Klaus Nielsen (Animal Disease Institute, Nepean, Ontario, and CAN). We strongly encourage other investigators to contribute their hybridomas to the CVB in the same spirit as did Klaus. However, it is important that the products of such hybridomas first be evaluated by competent labs for their specificity. Considering the small royalty generated by the sale of hybridomas to commercial suppliers, the tiny profit obtained after subtracting the cost of production and quality control, why not simply make them available free to other investigators through the CVB system. There are two exceptions to such a scheme: (a) if such hybridomas are the basis of “kits” sold by biotech reagent suppliers and (b) if the mAbs are for commercial use, not for investigators at universities and research institutes (click on REAGENTS to review the policy). Those who are considering contributing hybridomas should contact either Rick DeWald (rdewald@aphis.usda.gov) at the NVSL or J.E. Butler (john-butler@uiowa.edu) at the University of Iowa.
A second initiative is to publish lists of available mAb and the results of specificity tests on the CIgW Website (click on REAGENTS). An example is the soon to be published report evaluating the specificity of six anti-porcine IgA mAbs in regard to allotype bias/specificity. It is also important to report the mouse isotype of such mAbs since this can determine their usefulness in flow cytometry when dual labeling of lymphocytes is needed. The report on IgA mAbs will be followed by a preliminary report on the specificity of currently available anti-porcine IgG mAbs.
A third initiative is the preparation of a new generation of anti-IgG mAbs. In the case of swine, this is being funded through an NPB Toolkit grant which supports the preparation of heavy porcine-camelid chimeric subclass proteins for use in the preparation of mAbs and as reference standards for testing the specificity of existing mAbs. There is also a USDA-NRI Toolkit grant that funds the preparation of mAbs to a variety of Igs and antigens of interests in veterinary immunology.
Polyclonal reagents from commercial sources or raised by investigators continue to serve the veterinary community in various assays. However their shortcomings include: (a) their generally unproven or contested bias for subisotypes in all species, (b) background problems which they cause in immunohistochemistry and flow cytometry and (c) their lack of immortality. The latter has consequences for data reproducibility when certain polyclonals are replaced by others as supplies runs out. This can be especially problematic since there are no industry-wide standards in place and a number of labs make their own and have their own established internal reference standards. The reference standard issue could be managed through the Toolkit Workshop process. Therefore a future goal of “Toolkit Workshops” should be to reach agreement on universal standards that could potentially be supplied by the CVB. The next planned workshops are to be held in conjunction with the 9th International Veterinary Conference in Japan (contact Victor Rutten; V.rutten@vet.uu.nl) in 2010 and the 12th International Meeting of the Society for Developmental and Comparative Immunology in Prague in June 2009 (contact Imre Kacskovics;ikacsko@iif.hu).