AM&ku ChimicaActa, 281(1993) 13-18 Elsevier Science Publishers B.V., Amsterdam Detection of human T-lymphocytes with a piezoelectric immunosensor Bemd K&rig and Michael Gritzel Institut de Chimie Physique II, Ecole Polytechnique F&i&ale de Lausanne, CH-1015 Lausanne (Switzerland) (Received 26th January 1993; revised manuscript received 9th March 1993) A reusable pieaoelectric immunosensor was developed for the detection of purified human T-lymphocytes and of T-lymphocytes in whole human blood. Three different methods for immobilization of anti CD 3 gp M, 19 000-29OOfl antibody on the gold electrode of the immunosensor were tested. Coating the electrode with polyethyleneimine gave the best results in terms of stability, sensitivity and reproducibility. Similar results were observed on coating the electrode with y-aminopropyltriethoxysilane. The last method, coating the electrode with protein A, gave inferior results. Applying an anti CD 3 gp Mr 19OW-29000 antibody layer via polyethylenehnine immobilization on a lO-MHZ AT-cut crystal resulted in 5 X lo345 X 10’ cells on the surface in a linear frequency change and a long-term stability of 10 weeks when the modified crystal was stored desiccated at room temperature. Keyrvordr: Biosensors; Immunoassay; Piezoelectric sensing; Blood, Lymphocytes; T-lymphocytes The cellular and humoral immune reactions of mammalia are mediated by lymphocytes with a common bone marrow stem cell. In early child- hood two main groups of lymphocytes are gener- ated, the B-lymphocytes (bursa equivalent prolif- eration) and the T-lymphocytes (thymus-depen- dent proliferation). T-lymphocytes are responsi- ble for the celhdar immune response. They are found in the cortex of lymph nodes and are well characterized by different in vitro tests, e.g., T- cells can be stimulated by specific antigens such as tuberculin, a common proof in clinical diagnos- tics. A subpopulation of the T-cells, the so-called T-helper cells, cooperate with B-lymphocytes and induces the production of antibodies. This very important cell type is destroyed by the HIV virus. Another subpopulation of T-lymphocytes is the Correspondenceto: B. Kiinig, Institut de Chimie Physique II, Ecole Polytechnique FkdCrale de Lausanne, CT-I-1015 Lau- same (Switzerland). memory cells. They remember antigens and speed up the immune response during a second infec- tion with the same antigen. Other types are the T-suppressor and the T-effector cells; the latter synthesize lymphokines such as interferon, inter-’ leucin and the macrophage-inhibiting factor. Because T-cells play an important role in the immune system, it is of great interest to deter- mine the number of these cells per unit volume of whole blood. This is generally done by flow cy- tometry. This technique has two disadvantages, the high costs and the necessity to label the cells before use. A possible alternative is the use of piezoelectric antibody-based biosensors. Antibody-based biosensors detect the antigen concentration either by indirect competitive and displacement reactions similar to immunoassays or by direct changes in transducer output [l]. An example of the latter type of system is the piezo- electric (PZ) crystal detector for assays both in the gas phase and in solution. PZ devices consist ooO3-2670/93/$06.00 0 1993 - Elsevier Science Publishers B.V. AU rights reserved