Single mammalian cells compensate for differences in cellular volume and DNA copy number through independent global transcriptional mechanisms.

Individual mammalian cells exhibit large variability in cellular volume, even with the same absolute DNA content, and so must compensate for differences in DNA concentration in order to maintain constant concentration of gene expression products. Using single-molecule counting and computational image analysis, we show that transcript abundance correlates with cellular volume at the single-cell level due to increased global transcription in larger cells. Cell fusion experiments establish that increased cellular content itself can directly increase transcription. Quantitative analysis shows that this mechanism measures the ratio of cellular volume to DNA content, most likely through sequestration of a transcriptional factor to DNA. Analysis of transcriptional bursts reveals a separate mechanism for gene dosage compensation after DNA replication that enables proper transcriptional output during early and late S phase. Our results provide a framework for quantitatively understanding the relationships among DNA content, cell size, and gene expression variability in single cells.

C-Arm Fluoroscope Angle Settings for Fluoroscopically Guided Lumbar Transforaminal Epidural Injections.

BACKGROUND DATA: Minimizing fluoroscopy time in spine interventions is critical for time of procedure as well as radiation safety of the patient and medical personnel. Specific fluoroscopy angle settings for fluoroscopically guided L4-S1 transforaminal epidural injections (TFEIs) have not been described. OBJECTIVES: To describe the most common encountered settings for the C-arm fluoroscope angles for fluoroscopically guided L4-S1 (TFEI). METHODS: Each subject was placed in prone position on a flat fluoroscopy table without utilizing any device to alter innate lumbar spine curvature. The data from 246 consecutive patients at their first encounter in the fluoroscopy suite for a single level subpedicular lumbosacral TFEI was retrospectively analyzed. Most procedures occurred at the L4-5, L5-S1, and S1 levels (227 subjects). The C-arm angles including the oblique, cephalad/caudal were recorded for each subject upon observing final needle positioning for successful completion of the procedure according to ISIS Guidelines. RESULTS: For the L4-5 level, 71% of cases had oblique angle of 30°±5° and 94% of cases had neutral cephalad/caudal tilt (0°±5°) observed. For the L5-S1, 72% of cases had oblique angle of 30°±5° and 62% of cases had cephalad tilt angle of 15°±5° observed. For the S1 level, 73% of cases had oblique angle of 5°±5° and 69% of cases had cephalad tilt angle of 15°±5° observed. DISCUSSION/CONCLUSION: This retrospective descriptive study suggests fluoroscope angles for L4-S1 TFEI as a starting point before fine tuning views accounting for individual anatomy. Angles suggested for each level (oblique/cephalad tilt angles) are as follows: L4-5 (30/0°), L5-S1 (30/15°), and S1 (5/15°). Prospective studies using these guidelines would need to be undertaken to prove reproducibility between interventionalists, time efficiency, and radiation exposure reduction.

Multiplexed detection of viral infections using rapid in situ RNA analysis on a chip.

Viral infections are a major cause of human disease, but many require molecular assays for conclusive diagnosis. Current assays typically rely on RT-PCR or ELISA; however, these tests often have limited speed, sensitivity or specificity. Here, we demonstrate that rapid RNA FISH is a viable alternative method that could improve upon these limitations. We describe a platform beginning with software to generate RNA FISH probes both for distinguishing related strains of virus (even those different by a single base) and for capturing large numbers of strains simultaneously. Next, we present a simple fluidic device for reliably performing RNA FISH assays in an automated fashion. Finally, we describe an automated image processing pipeline to robustly identify uninfected and infected samples. Together, our results establish RNA FISH as a methodology with potential for viral point-of-care diagnostics.