About


Nile Red-stained vesicles in A549 cells (📷 B. Hariri)
We're an academic research lab located in the Department of Otorhinolaryngology at the University of Pennsylvania School of Medicine in Philadelphia.  

We study how airway cells sense and respond to infections.  Our major focus is on the nose and sinuses, also known as the upper airway.

Chronic upper respiratory infections can result in chronic rhinosinusitis (CRS), a disease affecting 8-10% of the US population with direct healthcare costs of over 6 billion dollars annually.  CRS has a major impact on quality of life, with CRS patients often suffering debilitating headaches from congestion and sinus pressure, a lack of ability to smell, and other detrimental symptoms.  CRS also has a major public health impact, as it accounts for 1 out of every 5 antibiotic prescriptions in adults in the US, making CRS treatment a major contributor to the emergence of antibiotic-resistant organisms.  A continuing goal of our research is to better understand airway physiology to identify new and better therapies to treat CRS and other airway diseases without the use of antibiotics.    


Another focus of the lab is cystic fibrosis (CF), the most common lethal recessive genetic disease in the US.  CF is characterized by altered fluid homeostasis in the lung, resulting in impaired mucociliary clearance and innate defense.  Our goal is to better understand the molecular and cellular bases of CF to help elucidate how to better treat the disease.

The tools we use include molecular biology, biochemistry, electrophysiology, and microbiology.  We specialize in using live-cell fluorescence and brightfield imaging to monitor real time cell function (physiology) in minimally invasive ways.

Research Support:

This lab has been generously supported by both private and federal grant funds.  


Current:

  • National Institutes of Health (NIH) National Institute for Deafness and Other Communication Disorders (R03DC013862, R01DC016309)
  • NIH National Institute of Allergy and Infectious Diseases (R21AI137484)
  • Cystic Fibrosis Foundation (LEER16G0)

Completed:

  • Start-up grant from the Department of Otorhinolaryngology-Head and Neck Surgery of the University of Pennsylvania Perelman School of Medicine, 
  • Pilot grant from the Penn Diabetes Research Center (DRC; funded through NIH DK19525)

Resources and Equipment:

General Molecular Biology, Biochemistry, and Cell Biology Equipment

  • Housed in BRB 1222-1223 (583 net square feet) and cold room 1263 (35 net square feet dedicated to our lab)
  • Centrifuges: Fisher Marathon Micro A, Eppendorf 5415C, Eppendrof 5402 refrigerated, Sorvall ST-8, Savant Speed Vac
  • General: Heatblocks, stir plates, balances, shakers, water bath, vortexers, etc.
  • DNA and protein electrophoresis and western blotting equipment 
  • Genesys UV/Vis spectrophotometer 
  • BioRad MyiQ single color real-time PCR detection system
  • BioRad GelDoc Imaging system
  • 4°C refrigerators, -20°C and -80°C freezers
  • Thermo Scientific MaxQ4450 incubated shaker
  • VWR Incu-Line and Fisher Isotemp bacteria incubators
Mammalian Cell Culture Incubators and Equipment
  • Housed in shared tissue culture suite 1274 (61 net square feet dedicated to our lab)
  • Thermo Scientific Forma 370 Stericycle CO2 mammalian cell incubators
  • Forma Tissue Culture Hood
Olympus Live-Cell Imaging System

 

  • Custom configured in collaboration with Olympus.  Housed in dedicated microscopy suite 1262 (99 net square feet)
  • Olympus IX-83 two-deck inverted microscope
  • 4x, 10x, 20x, 30x (silicone), 40x (LWD), 60x (oil) objectives
  • Differential interference contrast
  • Z-drift compensation (ZDC) laser focusing system (Olympus)
  • Lambda 10-2 dual filter wheels for ratiometric imaging with Smart Shutter (Sutter Instruments)
  • Filters for DAPI/SPQ, Fura-2, BCECF, FITC/GFP/Fluo-4/DAF-FM, TRITC/Cy3, CFP/YFP FRET/cameleon, SNARF (Chroma Technologies) 
  • Lambda LS 300 W Xe arc lamp (Sutter Instruments)
  • XCite LED 120 Boost LED light source
  • Olympus DSU spinning disk confocal with filters for DAPI, FITC, TRITC, Cy5 (Semrock), SNARF dual emission, and GFP/mCherry FRET   
  • Automated programmable stage (Prior)
  • Hamamatsu ORCA-Flash4.0 sCMOS camera (2Kx2K; 30 fps at full frame)
  • Tokai Hit (WSKM-F1) temperature-controlled and CO2-controlled incubation system for long-term time-lapse imaging of multi-well plates, including Neco control software and objective heater
  • Vibration isolation table (TMC) 
  • Heated (TC-344C), 6-channel perfusion system (Warner Instruments) for 35 mm glass bottom dishes (RC-37F) and coverslips (RC-26) with pinch valves
  • Peristaltic pump for perfusion
  • Bioptechs Delta-T heated chamber with perfusion lid and objective heater  
  • Imaging acquisition and analysis workstation (Dell Precision 7910 with Intel Xeon E5-2650 v3 processor, 32 GB RAM, 2x 512 GB SATA solid state drives, NVIDIA Quadro K2200 4GB GPU) with Metamorph Advanced for Olympus, MetaMorph NX 2.0, and MetaFluor for Olympus ratiometric imaging software, ImageJ, FIJI, Icy
  • Uses: Live-cell imaging of furs-2, fluo-4, DAF-FM, SNARF, BCECF, genetic GFP-based indicators, longer term imaging (5% CO2, 37 °C), 4-color immunofluorescence 
Nikon Imaging System
  • Housed in BRB 1262
  • Nikon TS100 inverted microscope
  • Phase contrast 10x and 100x objectives, 10x and 40x (LWD) PlanFluor objectives
  • 50W HBO Hg Arc Lamp
  • XCite 110 LED light source with instant on/off for shuttering
  • GFP/DAF-FM/Fluo-4 filter set
  • Photometric's Cool Snap HQ 12-bit cooled CCD camera
  • Color camera for histology
  • Micromanager (µManager) software for acquisition
  • Uses: Live cell imaging of GFP, calcium (fluo-4), nitric oxide (DAF-FM), etc.
TECAN Spark 10M Multi-Mode Plate Reader
  • Housed in BRB 1222
  • Absorbance
  • Top and bottom fluorescence with monochrometer excitation and monochrometer/filter fusion optics
  • High resolution area scan
  • Enhanced PMT
  • Incubation heating with carbon dioxide and oxygen control (including hypoxia) for real-time, live cell measurements
  • Two channel injection system
  • NanoQuant Plate for DNA/RNA quantification
  • SparkControl Magellan software for kinetic analyses
  • Uses: Endpoint assays (ELISA); DNA-RNA quantification; Real-time, live cell imaging of proliferation (XTT), mitochondrial membrane potential (JC-1), H2O2 production (amplex red), calcium (fluo-4), etc.
Transepithelial Electrophysiology Equipment
  • Housed in BRB 1222
  • World Precision Instruments DVC-1000 and Warner EC-825 dual channel epithelial voltage clamp amplifiers
  • Various Ussing chambers and accessories