Use of Iodixanol Self-Generated Density Gradients to Enrich for Viable Urothelial Cells from Nonneurogenic and Neurogenic Bladder Tissue

Bruce, A. Gregory; Sangha, Namrata; Richmond, Allison; Johnson, Kenny; Jones, Sarah; Spencer, Thomas; Ludlow, John W.

doi:10.1089/ten.tec.2009.0012

Cited by 2 publications

(2 citation statements)

References 16 publications

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“…7,8,27 Four-step density gradients, optimized for each species, separated UNFX cells into five subpopulations, designated B1 to B5. Detailed gene array analysis of rodent cells revealed that the B2 subpopulation was enriched for tubular epithelial cells (see Supplementary Data available online at www.liebertonline.com/ ten).…”

Section: Isolation Of Defined Subpopulations From Established Culturementioning

confidence: 99%

Isolation, Characterization, and Expansion Methods for Defined Primary Renal Cell Populations from Rodent, Canine, and Human Normal and Diseased Kidneys

Presnell

Bruce

Wallace

et al. 2011

Tissue Engineering Part C: Methods

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Chronic kidney disease (CKD) is a global health problem; the growing gap between the number of patients awaiting transplant and organs actually transplanted highlights the need for new treatments to restore renal function. Regenerative medicine is a promising approach from which treatments for organ-level disorders (e.g., neurogenic bladder) have emerged and translated to clinics. Regenerative templates, composed of biodegradable material and autologous cells, isolated and expanded ex vivo, stimulate native-like organ tissue regeneration after implantation. A critical step for extending this strategy from bladder to kidney is the ability to isolate, characterize, and expand functional renal cells with therapeutic potential from diseased tissue. In this study, we developed methods that yield distinct subpopulations of primary kidney cells that are compatible with process development and scale-up. These methods were translated to rodent, large mammal, and human kidneys, and then to rodent and human tissues with advanced CKD. Comparative in vitro studies demonstrated that phenotype and key functional attributes were retained consistently in ex vivo cultures regardless of species or disease state, suggesting that autologous sourcing of cells that contribute to in situ kidney regeneration after injury is feasible, even with biopsies from patients with advanced CKD.

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Section: Isolation Of Defined Subpopulations From Established Culturementioning

confidence: 99%

Isolation, Characterization, and Expansion Methods for Defined Primary Renal Cell Populations from Rodent, Canine, and Human Normal and Diseased Kidneys

Presnell

Bruce

Wallace

et al. 2011

Tissue Engineering Part C: Methods

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show abstract

“…To prevent sedimentation, a density-matching solution, such as OptiPrep (iodixanol) 89 , can be added to the growth medium in order to increase its density so that it matches that of the cells. Proper density matching prevents sedimentation and clumping, enabling encapsulation efficiency of beads and cells to remain nearly constant over periods of up to 2 h, without adverse cytotoxic effects 90,91 . An alternative method for preventing cell sedimentation and aggregation is to place a mini stir bar in the cell suspension syringe 27,92 .…”

Section: Introductionmentioning

confidence: 99%

Single-cell analysis and sorting using droplet-based microfluidics

et al. 2013

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We present a droplet-based microfluidics protocol for high-throughput analysis and sorting of single cells. compartmentalization of single cells in droplets enables the analysis of proteins released from or secreted by cells, thereby overcoming one of the major limitations of traditional flow cytometry and fluorescence-activated cell sorting. as an example of this approach, we detail a binding assay for detecting antibodies secreted from single mouse hybridoma cells. secreted antibodies are detected after only 15 min by co-compartmentalizing single mouse hybridoma cells, a fluorescent probe and single beads coated with anti-mouse IgG antibodies in 50-pl droplets. the beads capture the secreted antibodies and, when the captured antibodies bind to the probe, the fluorescence becomes localized on the beads, generating a clearly distinguishable fluorescence signal that enables droplet sorting at ~200 Hz as well as cell enrichment. the microfluidic system described is easily adapted for screening other intracellular, cell-surface or secreted proteins and for quantifying catalytic or regulatory activities. In order to screen ~1 million cells, the microfluidic operations require 2–6 h; the entire process, including preparation of microfluidic devices and mammalian cells, requires 5–7 d.

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Use of Iodixanol Self-Generated Density Gradients to Enrich for Viable Urothelial Cells from Nonneurogenic and Neurogenic Bladder Tissue

Cited by 2 publications

References 16 publications

Isolation, Characterization, and Expansion Methods for Defined Primary Renal Cell Populations from Rodent, Canine, and Human Normal and Diseased Kidneys

Isolation, Characterization, and Expansion Methods for Defined Primary Renal Cell Populations from Rodent, Canine, and Human Normal and Diseased Kidneys

Single-cell analysis and sorting using droplet-based microfluidics

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