Plastic membranes containing deoxyribonucleic acid (DNA) as an electroactive material were acting as Ca2+ selective sensors. Diethyl phthalate (DEP), dioctyl Phthalate (DOP), or nitrophenyl octyl ether (NPOE) were used as plasticizers and polyvinyl chloride (PVC) was the membrane matrix. A sensor with a membrane composition of 120 mg PVC, 60 mg DOP plasticizer, and 2 mg DNA ionophore (DNA: DOP: PVC, 1.0:29.2:0.1 mole) was found to have the best performance. The slope of the calibration graph was 30 mV decade􀀀1. The optimum pH range was 5.7–9.5 for 0.01 M Ca2+. The sensor response time was fast (2–3 s) with a long working period (up to 3 weeks). Excellent selectivity for Ca2+ was indicated by the values of selectivity coefficients for different selected interference. The sensor was used effectively for the estimation of calcium in real samples (fruits, calcium syrup, milk, and dairy products).