Automated monitoring of cell concentration and viability using an image analysis system

Abstract

In order to automate measurements of cell concentration and viability in a suspended animal cell culture, we have developed an in situ microscopic image analysis system with an effective cell recognition algorithm. With a small amount of sample, this system can measure the cell density rapidly and aseptically. In addition, it can measure a cell size histogram including cell debris small particle distribution. These small particles have been found to be related to the viability of the mouse-mouse hybridoma STK1 cell line. By using cell debris small particle density as an indicator of cell viability, the developed system provides non-destructive viability monitoring without trypan blue staining.

Validation of three viable-cell counting methods: Manual, semi-automated, and automated

“…This method depends on the analyst’s ability to evaluate different cell attributes regardless of the cell type; in addition allows using different staining techniques according to the purpose of the analyses ( Table 1 ). However, the procedure is time consuming, which precludes the analysis of a large amount of samples at one time and is subject to inter-user variation depending on the degree of expertise of the analyst [6] , [8] .…”

“…In general, automated cell count instruments consist of a digital camera to obtain images and the analyses are performed through specialized software that requires a minimal user involvement [10] . Although automated instruments facilitate the process of analyzing samples, they are constrained by the availability of a few compatible staining options, and may be imprecise in differentiating some types of cells due to technical limitations in their hardware and software [12] , [8] .…”

Biomass Quantification by Image Analysis

“…On-line systems involve circulation loops to bring the sample to the microscope. The depth-of-focus is limited and the cells should be maintained close to the focus plane by forcing them through a capillary [38] or a flow cell [39]. For Bacillus thuriengiensis, the best results were obtained by Lichtfield et al [38] with 20 mm thick glass channels.…”

“…The dead cells are lysed and their intracellular content is dispersed in the broth [166]. Automated analysis of hybridoma cell viability has been proposed by Tucker et al [167] and Maruhashi et al [39] using Trypan Blue staining.…”

Optical analysis of perfusion bioreactor cell concentration in an acoustic separator

“…Optical turbidity sensors (Konstantinov et al, 1992;Zhou and Hu, 1994), have been an important alternative to sampling and off-line analysis (Ulber et al, 2003;Wu et al, 1995). Also, microscopic image analysis can be used (Maruhashi et al, 1994). These optical systems often become unreliable due to bubbles or fouling of the probes and may require replacement during bioreactor operation (Matanguihan et al, 1999).…”