Check GAD antibody positivity with the Diamyd anti-GAD RIA plate*

100GAD in GraphsGAD65 DNA vaccination preventsdiabetes in NOD micediabetic mice (in percent)90ppInsulin (n=25)80 ICA69+GAD65+ppIns (n=14)control (untreated) (n=14)70 control (vector) (n=12)GAD65 (n=14)6050403020DNA vaccination1000 10 20 30age (in weeks)Karges et al, Diabetes 51:3237; 2002Proliferation (cpm)• 4-6 week old female NOD• Different DNA vaccinations• Insulin construct does not protectfrom diabetes• GAD65 construct preventsdiabetes• Combining constructs abolishesGAD65-induced protectionBaculovirus-encoded recombinantGAD outperforms bacterial andyeast GAD preparations in T cellproliferation assays100000100001000baculovirusyeast100bacterial1 10Peakman et al (2002) Characterization of preparationsof GAD65, proinsulin and IA-2 for use indetection of autoreactive T-celsl in Type 1 diabetes.Diabetes 50:1749-1754• T cell recall immune responseswere studied by proliferation assaysusing defined T cell lines and clonesand different preparations of GAD65• Two independently producedGAD65 preparations expressed bybaculovirus gave higher proliferationindices than either yeast- orbacterial-produced GAD65Untangling the GADsAllan TobinMy interest in GABA and GAD came from my interest in Huntington’s disease – adegenerative neurological disorder that had killed the mother of two of my friends.I decided in the early 1980s that GAD was a reasonable “candidate gene” forHuntington’s disease, and I set out to isolate it.The time was ripe for gene isolationsince new molecular techniques were emerging every month – both new ways ofmaking collections of genes and new methods for screening them for genes of interest.Just at that time, Dan Kaufman, whohad worked in my laboratory as aUCLA undergraduate and thenmoved to Berkeley to do graduatework, decided to move back to LosAngeles, to rejoin my laboratory, andto work on the brain.Never suspecting that I would everend up contributing to that field, I counseled Danto work on a more tractable problem – the isolationof the GAD gene. At the time, few neuroscientistswere using molecular techniques, and findingthat gene would undoubtedly make a significantcontribution to the field. Even ifHuntington’s disease is not caused by a mutationin GAD, I reasoned, having the gene in handwould accelerate progress in understanding theinhibitory circuits that are important in normalbrain function and that fail in Huntington’s disease.Dan quickly succeeded in finding the GAD gene(which we now call GAD67), using a combinationof immunological, molecular, and biochemical techniques.Pure luck – the ability of bacteria to makea functional GAD – allowed us to short circuitmost of the slogging that we thought would benecessary to prove we had the right gene. But therewere some anomalies that later caused another graduatestudent, Mark Erlander, to challenge our initialview that “there is only one GAD”. Using a newset of techniques, in the early days of PCR (thepolymerase chain reaction), Mark found anotherGAD gene, which we now call GAD65.In 1990, Dan Kaufman, then a postdoctoral felllowat the Salk Institute, again returned to thelab, again interested in autoimmune disease. Onthe basis of a paper by Pietro Di Camilli on StiffMan Syndrome – a rare complication of diabetesand of other conditions, Dan suspected thatGAD65 or GAD67 might be identical to theunnamed 64 kD antigen first identified by ÅkeLernmark in 1982 as the first target of Type 1(T1D) antibodies. About the same time, SteinunnBaekkeskov, Pietro Di Camilli, and their colleaguescame to a similar conclusion though they didn’tknow about GAD65, which we had onlyrecently discovered. In collaboration with NoelMaclaren and Mark Atkinson at the Universityof Florida, we examined GAD autoimmunity inT1D patients, using GAD65 and GAD67 that wecould produce from recombinant DNA.Autoantibodies to GAD indeed provided anpage 14dmccad june 2003