GSM2299958: AV macrophage, Transcriptome 8; Mus musculus; RNA-Seq
AV node macrophages were FACS-purified from whole AV node tissue as described above. Single macrophages were then captured using the Fluidigm C1 microfluidic chip designed for 5- to 10-µm cells according to the manufacturer’s protocol. A concentration of 1.8x105 cells per ml was used for chip loading. After cell capture, chips were examined visually to identify empty chambers, which were excluded from later analysis. Cell lysis and cDNA synthesis were performed on-chip with Clontech SMARTer Ultra Low RNA kit for the Fluidigm C1 system. Amplified cDNA was validated and quantified on an Agilent Bioanalyzer with the High Sensitivity DNA chip. Illumina libraries were constructed in 96-well plates using the Illumina Nextera XT DNA Sample Preparation kit according to a modified protocol supplied by Fluidigm. Constructed libraries were validated and quantified with the Agilent High Sensitivity DNA chip, and subsequently normalized and pooled to equal concentrations. The pooled libraries were quantified by qPCR using KAPA Library Quantification kit (Kapa Biosystems).