Serum-induced basophil CD63 expression by means of a tricolour flow cytometric method for the in vitro diagnosis of chronic urticaria.
BACKGROUND
Functional autoantibodies against the alpha-chain of the high-affinity IgE receptor (FcepsilonRIalpha) identify a subset of patients with chronic urticaria (CU) due to autoreactivity, as assessed by an in vivo positive response to autologous serum skin test (ASST). We performed a study to standardize the serum-induced basophil activation assay by flow cytometry (FCM) using a new tricolour method, assessing the diagnostic performance of this test in discriminating between ASST+ and ASST- CU patients.


METHODS
Sera of 64 CU patients (22 ASST+ CU and 42 ASST- CU) and 10 healthy subjects were tested for their ability to induce basophil CD63 expression when incubated with whole blood of both atopic (DA) and non-atopic donors (DNA). Using a triple-labelled strategy with anti-CD123, anti-HLA-DR and anti-CD63 antibodies, CD63+ basophils were identified on a selected population of CD123+ HLA-DR- cells. In 3 ASST+ CU patients who underwent cyclosporine therapy, the assay was performed before and after treatment.


RESULTS
The ASST+ CU sera resulted in a significant higher induction of basophil CD63 expression compared with ASST- CU and healthy donors sera; when whole blood from DA was used, sensitivity and specificity of the assay were 95.5% and 90.5% respectively. ASST+ CU serum activity was significantly decreased during cyclosporine A treatment, in parallel with clinical remission.


CONCLUSIONS
Chronic urticaria serum-induced CD63 expression assay performed on DA whole blood by means of our tricolour FCM method could be the most useful tool for identification of a subset of patients with autoimmune CU and may become a promising tool also for monitoring treatment efficacy.
Frezzolini
A
A
Laboratory of Immunology and Allergology, Istituto Dermopatico dell'Immacolata, IDI-IRCCS, Rome, Italy.
Provini
A
A
Teofoli
P
P
Pomponi
D
D
De Pità
O
O
D000328:Adult; D015703:Antigens, CD / Q000096:biosynthesis / Q000097:blood / Q000235:genetics*; D001491:Basophils / Q000276:immunology / Q000378:metabolism*; D002469:Cell Separation; D002908:Chronic Disease; D005260:Female; D005434:Flow Cytometry / Q000379:methods; D006801:Humans; D006969:Hypersensitivity, Immediate / Q000097:blood / Q000276:immunology; D008297:Male; D008875:Middle Aged; D010980:Platelet Membrane Glycoproteins / Q000096:biosynthesis / Q000235:genetics*; D012680:Sensitivity and Specificity; D044967:Serum / Q000276:immunology*; D060149:Tetraspanin 30; D014581:Urticaria / Q000097:blood / Q000175:diagnosis* / Q000276:immunology*
D015703:Antigens, CD; C554586:CD63 protein, human; D010980:Platelet Membrane Glycoproteins; D060149:Tetraspanin 30