Nanoparticles are widely used to separate and detect bacteria by immunoassay techniques. However, there is a great need to develop a new low cost, easy-handling, and fast bacteria quantification method in order to give fast response to patients, when there are limited time and instruments. This article describes a new nanoparticle-based quantification method by using only sucrose gradient centrifugation and an easy optical setup. First of all, spherical-shaped nanoparticles with different chemical components have been synthesized. Buoyancy tendencies of these nanoparticles and effects of different mediums were examined to obtain moving particle band which is necessary for rate-zonal centrifugation. Optimum gradient and process parameters were determined; then, moving bands of nanoparticle and bacteria-conjugated nanoparticle were analyzed by a software. Migration distance of bacteria-captured nanoparticle bands was found inversely proportional to bacterial concentration. Bacteria-nanoparticle conjugates were characterized by transmission electron microscopy images and zeta potential measurements. The developed method enables non-specific detection and quantification of E. coli K-12 within the range of 10(5)-10(8) cfu/ml, by using chitosan-coated CdTe quantum dots. Chitosan-coated CdTe quantum dots were found advantageous for the easy and fast tracking of bacteria-particle conjugate bands in the developed gradient method with respect to their high bacterial capture, sedimentation rate, and light emission properties. According to our findings, the proposed gradient method was found to be an advantageous bacteria quantification method with limited instrument requirement and 50 min of total detection time in solution.