Unit 1 · Basics in cell culture (Weeks 1 to 5)
Week 1: Intro to Cell Culture and the Cx43 Research Story
Course logistics and final project, history and uses of cell culture, and her gap junction (Cx43) research in pancreatic cancer cells.
Week 2: Observing Cells and Aseptic Technique
Types of cells in culture, cell health and mitosis, contamination (bacteria, mycoplasma) and aseptic technique.
Week 3: Senescence, Immortalization and Subculture
Immortal cell sources, senescence and telomerase, immortality vs neoplasia, subculture by enzymatic dissociation, and the hemocytometer slides.
Week 4: Cell Quantitation
Accuracy and subculture, quantitation of cells, cells per flask and seeding density.
Week 5: Cryopreservation
Freezing damage, supercooling, dehydration, vitrification, DMSO and storage. The updated Sep 24 deck (replaces the first cryo deck).
Week 5: Vitrification Paper
Oocyte vitrification paper (slides 1 to 14). Slides 15 to 27 repeat the Microscopy deck.
Week 5: Microscopy
Light microscopy, resolution, electron microscopy, and use and care of the microscope.
Unit 2 · Cellular assays (Weeks 6 to 8)
Week 6: Stains and Fluorescence
Cellular stains, fluorescence (excitation and emission, GFP, FRAP, FRET, flow cytometry, immunofluorescence), DAPI and phalloidin, wide field vs confocal.
Week 7: Cell Proliferation, Trypan Blue and Research Papers
Growth curves and population doubling time, mitotic index, plating efficiency, viability, IC50 by trypan blue, and multi-well plates.
Week 8: Cytotoxicity and Survival Curves
Cytotoxicity, in vitro limits, cytotoxicity assays (ATP, Alamar Blue redox), survival curves and IC50.
Unit 3 · Cytotoxicity projects and qPCR (Weeks 9 to 13, posted ahead)
Week 9: Primer Design (Intro to qPCR)
Central dogma, PCR and qRT-PCR, and primer design criteria for the final project.
Week 10: Apoptosis and Neoplasia
Drug response: apoptosis (caspases, Bcl-2, intrinsic and extrinsic pathways, detection assays), neoplasia and cell signaling.
Weeks 11 to 13: qPCR Experiment (RNA Isolation to cDNA)
RNA isolation (RNeasy), reverse transcription to cDNA, and amplifying with qPCR.
Week 13: qPCR Experiment and Analysis
Dilution series, R2, efficiency, melting curve, absolute vs relative quantification, controls and the qPCR reaction.