A flexible and cost-effective manual droplet operation platform for miniaturized cell assays and single cell analysis.

Herein, we developed a flexible and cost-effective manual droplet operation system (MDOS) for performing miniaturized cell assays as well as single cell analysis. The MDOS consists of a manual x-y-z translation stage for liquid transferring and switching, a high-precision syringe pump for liquid driving and metering, a tapered capillary probe for droplet manipulation, a droplet array chip for droplet loading and reaction, sample/reagent reservoirs for storage, and a microscope for droplet observation, with a total expense of only $4,000. By using the flexible combination of three elementary operations of the x-y-z stage's moving and the pump's aspirating and depositing, the MDOS can manually achieve multiple droplet handling operations in the nanoliter to picoliter range, including droplet generation, assembling, fusion, diluting, and splitting.

A Microfluidic Droplet Array System for Cell-Based Drug Combination Screening.

In the last few decades, drug combination therapy has been widely applied in oncology and in other complex diseases. Due to its potential advantage of lower drug toxicity and higher therapeutic efficacy, drug combination treatment has been more and more studied in fundamental labs and pharmacy companies. In this chapter, we report cell-based drug combination screening using a microfluidic droplet system based on a sequential operation droplet array (SODA) technique.

Three-Dimensional Cell Culture and Drug Testing in a Microfluidic Sidewall-Attached Droplet Array.

Three-dimensional (3D) cell culture provides an effective way over conventional two-dimensional (2D) monolayer culture to more closely imitate the complex cellular organization, heterogeneity, and interactions as well as tissue microenvironments in vivo. Here we present a novel droplet-based 3D cell culture method by using droplet array attached on the sidewall of a PDMS piece. Such an arrangement not only avoids cells from adhering on the chip surface for achieving 3D cell culture in nanoliter-scale droplets, but also facilitates performing multiple operations to cells in droplets, including cell suspension droplet generation, drug treatment, and cell staining with a capillary-based liquid handling system, as well as in situ observation and direct scanning with a confocal laser scanning microscope.

Microdroplet chain array for cell migration assays.

Establishing cell migration assays in multiple different microenvironments is important in the study of tissue repair and regeneration, cancer progression, atherosclerosis, and arthritis. In this work, we developed a miniaturized and massive parallel microfluidic platform for multiple cell migration assays combining the traditional membrane-based cell migration technique and the droplet-based microfluidic technique. Nanoliter-scale droplets are flexibly assembled as building blocks based on a porous membrane to form microdroplet chains with diverse configurations for different assay modes.

Cell-based drug combination screening with a microfluidic droplet array system.

We performed cell-based drug combination screening using an integrated droplet-based microfluidic system based on the sequential operation droplet array (SODA) technique. In the system, a tapered capillary connected with a syringe pump was used for multistep droplet manipulations. An oil-covered two-dimensional droplet array chip fixed in an x-y-z translation stage was used as the platform for cell culture and analysis.

Hepatocellular carcinoma-associated protein markers investigated by MALDI-TOF MS.

Hepatocellular carcinoma (HCC) is one of the most common types of cancer worldwide. The initial hepatocellular alterations that precede the appearence of HCC include chronic viral hepatitis/cirrhosis, foci of phenotypically altered hepatocytes and, subsequently, dysplastic hepatocytes that form foci and nodules. These changes cause a discrepancy in the microenvironment of liver cells, which may result in changes in the protein expression profile of the cells.

[Effect of recombinant human erythropoietin on bcl-2 protein expression in the retina in a rabbit model of acute high intraocular pressure].

Objective: To investigate the effect of recombinant human erythropoietin (rhEPO) on the expression of bcl-2 protein in the retina of rabbits with acute high intraocular pressure and explore the mechanism underlying the protective effect of rhEPO on the retina against ischemia-reperfusion injury.

Methods: rhEPO was injected subcutaneously in the ear of a rabbit model of acute high intraocular pressure induced by physiological saline perfusion into the anterior chamber. Bcl-2 protein expression in the retina of the rabbits was observed by immunohistochemical staining on days 1, 3, 7, and 14 after retinal ischemia-reperfusion and compared with that in normal rabbits and untreated rabbit models.

[Effects of rAAV-mediated rhBDNF gene transfection on BDNF gene expression in the retina of a rabbit model of acute high intraocular pressure].

Objective: To observe the changes in the expression of brain derived neurotrophic factor (BDNF) gene in the retina of rabbits with acute high intraocular pressure (IOP) after injection of recombinant adeno-associated virus (rAAV) vector containing human BDNF gene (rAAV-hBDNF), and investigate the neuroprotective mechanism of rAAV-hBDNF.

Methods: The unilateral eyes of 24 white rabbits were randomly chosen as the model group with high IOP induced by saline perfusion into the anterior chamber, and the contralateral eyes served as the control group without treatment. In another 24 white rabbits, 10 microl rAAV-BDNF was injected into the vitreous body of one of the eyes 3 days before induction of high IOP.

[Neuroprotective effect of rAAV-mediated rhBDNF gene transfection on rabbit retina against acute high intraocular pressure].

Objective: To investigate the neuroprotective effect of human brain-derived neurotrophic factor gene transfection into rabbit retina against acute high intraocular pressure (HIOP).

Methods: Acute HIPO was induced in one eye of 24 white rabbits via saline perfusion into the anterior chamber (model group), and the contralateral eye without treatment served as the control group. In another 24 rabbits, 10 microl recombinant adeno-associated virus (rAAV) vector containing human BDNF gene (rAAV-BDNF) was injected into the vitreous body of one of the eyes 3 days before the operation for HIPO (BDNF group).

[Effects of recombinant human erythropoietin on hypoxia inducible factor-1alpha expression in the retina of rabbits with acute high intraocular pressure].

Objective: To observe the effect of recombinant human erythropoietin (rhEPO) on the expression of hypoxia inducible factor-1alpha (HIF-1alpha) in the retina of rabbits with acute high intraocular pressure and investigate the mechanism of rhEPO in protecting the retina from ischemia-reperfusion injury.

Methods: Acute high intraocular pressure was induced in the rabbits by perfusion of normal saline into the anterior chamber, and rhEPO was injected subcutaneously. The changes in HIF-1alpha protein expression in the retina was observed by immunohistochemistry on days 1, 3, 7, and 14 after retinal ischemia- reperfusion.

[Histomorphology and proteomics analysis of human fetus liver].

Objective: To investigate the histomorphology and protein expression profiles of human fetal liver at different developmental stages.

Methods: The protein expression patterns of human fetus livers at early and late stages were profiled by two-dimensional gel electrophoresis (2-DE) combined with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) analysis. The differentially expressed proteins were identified by searching the protein databases with Mascot or ProFound softwares.

[Study on tetrahydrobiopterin deficiency in Northern Chinese population].

Objective: To emphasize early differential diagnosis from patients with hyperphenylalaninemia (HPA) and to evaluate the treatment and long-term outcome of patients with tetrahydrobiopterin synthase (BH4) deficiency in Northern Chinese population.

Methods: From 1992 to 2005, a total of 618 patients with HPA were diagnosed and/or cared for in our outpatient clinic. Urinary pterin analysis, detection of dihydropteridine reductase (DHPR) activity in blood, and then BH4 loading tests were carried out to differentiate BH4 deficiency in these patients from classical phenylketonuria.