The effect of varying conditions to Testosterone-3-carboxymethyl oxime-4-(10-methy)-acridinium-9-carboxylate (To-3-AEPMAC) on the light yield and binding affinity of Testosterone chemiluminescent-marker conjugate was investigated. For this purpose To-3-AEPMAC has been synthesized

the resulting was then evaluted as potential labels for the development of an immunoassay based on monitoring chemiluminescence. The conjugate was able to compete with testosterone for the binding siles of an antibody raised against To-3-CMO bovine serum albumin

and they showed high affinity as same as unaltered testosterone. The conjugate produced light up on oxidation by a hydrogen peroxide in basic condition and was not significantly affected the quantum yield. The chemiluminescent reaction was optimized in terms of pH 6.0 concentration of oxidant

sodium hydrate and conjugate design. Under optimal conditions

the conjugate was detectable at femtomolar levels. The lowest detection limit was obtained using To-3-AEP-MAC (0.18fmol). These results indicated that conjugate does not modify its chemiluminescent property and immunoreativity

the immunoassay techniques based on chemiluminescence can be developed with the label.