Single-Cell Analysis: Methods and Protocols by Dino Di Carlo, Henry Tat Kwong Tse, Daniel R. Gossett

By Dino Di Carlo, Henry Tat Kwong Tse, Daniel R. Gossett (auth.), Sara Lindström, Helene Andersson-Svahn (eds.)

Today, cells are in general analyzed en masse, with hundreds of thousands of cells according to pattern, yielding effects at the general reaction of the cells. notwithstanding, mobile heterogeneity signifies that as a way to study extra approximately mobile behaviour, it is very important research how person cells reply, one after the other. In Single-Cell research: equipment and Protocols, specialists within the box offer an replace at the box of single-cell research in which the newest findings and purposes are defined intimately. The equipment defined during this publication contain a number of examples of traditional equipment and several other examples of miniaturized equipment. Written within the hugely winning Methods in Molecular Biology™ series structure, chapters contain introductions to their respective subject matters, lists of the mandatory fabrics and reagents, step by step, effortlessly reproducible laboratory protocols, and key tips about troubleshooting and keeping off recognized pitfalls.

Authoritative and available, Single-Cell research: equipment and Protocols encourages readers to discover new methods of learning cells which could aid bring about fascinating new discoveries.

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Distribute the liquid with a sterile cell scraper and remove excess liquid by scraping it over (the edge of) the plate. Each microwell should be compartmentalized from the neighboring well, containing a fixed volume of culture medium. 7. Place the microwell plate in the standard slide holder in a flow cytometer with plate-sorting capabilities. 8. Begin automatic single-cell seeding (time: approximately 1 well/s). , 1 cell) is sorted in each well (Fig. 3, day 1). The wells are already prefilled with growth medium (see step 6 above).

B. Brown et al. 19. Column tip is fragile, so try not to touch the actual tip with the tube/microwell to avoid breakage. 20. Do not touch the top surface of the cap if possible, any dirt/ oil will cause the small droplet to spread on the cap and lead to adsorption of sample. References 1. Arkhipov, S. , and Krylov, S. N. (2005) Chemical cytometry for monitoring metabolism of a Ras-mimicking substrate in single cells, Cytometry A 63, 41–47. 2. , Sims, C. , Stanbridge, E. , and Allbritton, N. L. (2004) A quantitative single-cell assay for protein kinase B reveals important insights into the biochemical behavior of an intracellular substrate peptide, Biochemistry 43, 1599–1608.

Andersson-Svahn Fig. 1. (a) Schematic side view of the microplate. The semipermeable membrane consists of PDMS and is manually added after cell seeding. The KOH etched silicon microgrid creates squared microwells when anodically bonded to the glass bottom. Each microwell has sloped walls with the dimensions 1,360 × 1,360 mm2 (top) and 650 × 650 mm2. The pitch between each well is 140 mm, giving a center–center distance of 1,500 mm between the wells. 5 ml. Autoclave at 121°C for 20 min and let the plates cool to room temperature.

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