Abstract: A series of assistant reagents were added to the horseradish peroxidase (HRP)/luminol/enhancer/H 2O 2 system, and got a new type of enhanced chemilumine- scent substrate solution catalyzed by HRP. It has the advantage of high sensitivity, slow decay of light emission, being stored for long time, and so on. The assay sensitivity for HRP is 2.4 amol per well. The half-life of decay of light emission is about 40 min. The work solution can be stored for more than 14 d at 37 ℃ or 30 d at room temperature. A two-site sandwich BAS ECLEIA for TSH and a two-site sandwich ECLEIA for AFP were developed. In the TSH-BAS ECLEIA, the range of detection is 0.17～35 mIU/L. The signal to background ratio is above 5.The assay sensitivity and function sensitivity are 0.04 mIU/L and 0.06 mIU/L, respectively. The intra-assay and inter-assay coefficients of variation are 2.96%～5.05% and 4.01%～7.56%, respectively. The recovery is 101%～119%. The correlation coefficient and the correlation equations of sample dilution are 0.996, y=44.65x+0.352. The correlation coefficient and the correlation equations of assay with Bayer ACS:180 system are 0.984, y= 0.889x-0.351. In the AFP ECLEIA, the range of detection is 5～2 000 ng/mL. The signal to background ratio is above 4. The kinetics experiment shows that the result is accurate and reliable in 15 min after initiation the chemiluminescent reaction.