microfluidic polydimethylsiloxane co-flow devices flowjem drop-seq chips  (FlowJEM Inc)

Journal: BMC Biology

Article Title: Cell fixation and preservation for droplet-based single-cell transcriptomics

doi: 10.1186/s12915-017-0383-5

Figure Lengend Snippet: Cell preparation for droplet-based single-cell transcriptional profiling. Schematic of experimental workflow. Cultured human ( HEK ) and mouse ( 3T3 ) cells were dissociated, mixed and further processed to analyse the transcriptomes of either live or fixed cells by Drop-seq. Washed cells were gently resuspended in 2 volumes of ice-cold PBS, then fixed by adding 8 volumes of ice-cold methanol. Methanol-fixed cells could be stored for up to several weeks at –80 °C. Prior to Drop-seq, cells were washed before passing them through a 35- to 40-μm cell strainer. Cells were then separately encapsulated in droplets together with a single bead in a microfluidic co-flow device and single-cell transcriptomes sequenced in a highly parallel manner. Downstream analysis and systematic quantitative comparisons were subsequently made from separate experiments using live or fixed cellular input material with an R package ('dropbead') that we developed and is freely available for download

Article Snippet: Monodisperse droplets of about 1 nl in size were generated using microfluidic polydimethylsiloxane (PDMS) co-flow devices (Drop-seq chips, FlowJEM, Toronto, Canada; rendered hydrophobic by pre-treatment with Aquapel).

Techniques: Cell Culture