BME 409 Key Terms for Exam 2

These flashcards cover key terms and concepts related to verification, validation, safety, microscopy techniques, immunology, and protein assays as discussed in BME 409 for exam preparation.

Verification

Confirming a method or device performs as intended.

Validation

Demonstrating that results are accurate, reliable, and appropriate for their intended use.

Safety and Efficacy

Ensuring a product is safe and produces the desired biological effect.

Characterization

Measuring physical, chemical, or biological properties of a material or sample.

Biocompatibility

The ability of a material to function in the body without causing harm.

In Vitro Studies

Experiments performed outside a living organism, typically in culture.

In Vivo Animal Studies

Testing performed in living animals to assess biological responses.

In Vivo Human Studies

Clinical studies evaluating safety and performance in humans.

Wide-Field Microscopy

Illuminates the entire sample; good for general imaging.

Confocal Microscopy

Uses point illumination and optical sectioning for high-resolution 3D imaging.

Electron Microscopy

High-magnification imaging using electron beams to visualize ultrastructure.

Cultured Cells

Cells grown and maintained in controlled laboratory conditions.

Biopsied Tissue

Tissue removed from a living organism for analysis.

Microtome

Instrument that slices thin tissue sections for microscopy.

Morphology

The shape, structure, and appearance of cells or tissues.

Confluence

The percentage of a culture dish covered by cells.

H&E Stain

Histology stain: hematoxylin (nuclei blue) and eosin (cytoplasm pink).

Hematocytometer

A counting chamber used to measure cell concentration.

Image Analysis

Quantifying features in images using software.

Destructive Assays

Tests that require destroying the sample.

Migration Chambers

Devices that measure cell movement across a membrane or gradient.

Fluorescence

Emission of light by a molecule after excitation.

Immunofluorescence

Using fluorescent antibodies to detect specific proteins.

Quantum Dots

Bright, stable fluorescent nanoparticles used for labeling.

Green Fluorescent Protein (GFP)

A naturally fluorescent protein used as a reporter.

Antibody

A protein produced by B cells that binds specific antigens.

Antigen

A molecule recognized by an antibody or immune receptor.

Epitopes

The specific part of an antigen that an antibody binds.

Paratope

The region of an antibody that binds the epitope.

Mimotope

A peptide that mimics an epitope’s structure.

B Lymphocytes

Immune cells that produce antibodies.

Monoclonal Antibodies

One epitope, one clone.

Polyclonal Antibodies

Multiple epitopes, multiple clones.

Primary Antibodies

Binds the target antigen.

Secondary Antibodies

Binds the primary antibody and carries a label.

Direct ELISA

Antigen is captured and detected with a labeled primary antibody.

Indirect ELISA

Antigen is captured, primary antibody binds, and a labeled secondary detects it.

SDS-PAGE

Gel electrophoresis that separates proteins by size.

Polyacrylamide Gel

The matrix used in SDS-PAGE to separate proteins.

Western Blotting / Immunoblotting

Proteins are transferred to a membrane and detected with antibodies.

What are the methods we use for evaluating a TEMP?

Verification, validation, biocompatibility testing, in vitro assays, in vivo animal studies, imaging, mechanical testing, and biochemical analysis.

What is the function of light microscopy in tissue engineering?

To visualize cells and tissue structure, assess morphology, confluence, and general health of engineered constructs.

How do we determine cell morphology from light microscopy?

By examining cell shape, size, spreading, alignment, and structural organization under bright‑field imaging

How do we determine cell proliferation from light microscopy?

By tracking increases in cell number, confluence changes, or using proliferation stains visible under light microscopy.

How do we determine cell migration from light microscopy?

By performing scratch assays or migration chamber imaging and tracking cell movement over time.

How do we determine cell apoptosis from light microscopy?

By identifying morphological hallmarks such as cell shrinkage, blebbing, fragmentation, or using apoptosis-specific stains.

How do we determine cell differentiation from light microscopy?

By observing changes in morphology, staining patterns, or marker expression visible with histological or immunostaining techniques.

How do we use fluorescence to identify tissue components?

Fluorescent dyes or tagged antibodies bind specific molecules, allowing visualization of proteins, nuclei, or structures under fluorescence microscopy

What is the natural function of antibodies? What is their structure?

Antibodies recognize and bind antigens to neutralize pathogens. They have two heavy and two light chains forming variable antigen‑binding regions

What is the difference between monoclonal and polyclonal antibodies?

Monoclonal antibodies bind one epitope from a single B‑cell clone; polyclonal antibodies bind multiple epitopes from multiple clones

How do we use antibodies to detect a specific molecule?

By binding the antibody to the target molecule and visualizing it using fluorescence, enzymes, or secondary antibodies.

How do we detect and analyze specific proteins in tissues?

Using immunohistochemistry, immunofluorescence, Western blotting, or ELISA to identify and quantify protein expression.

Genomics

Study of the complete DNA sequence of an organism.

Transcriptomics

Analysis of all RNA transcripts produced by cells.

Proteomics

Study of all proteins expressed in a cell or tissue.

Metabolomics

Measurement of small‑molecule metabolites in a system.

Phenomics

Comprehensive study of observable traits and behaviors.

Nucleotides — ATGC

The four DNA bases: adenine, thymine, guanine, cytosine.

Hybridize

To bind complementary DNA or RNA strands together.

Sanger sequencing

DNA sequencing using chain‑terminating ddNTPs.

dNTP vs ddNTP

dNTP: normal nucleotide;

ddNTP: lacks 3’ OH, stops DNA synthesis.

DNA polymerase

Enzyme that synthesizes DNA from a template strand.

Polymerase Chain Reaction (PCR)

Technique to amplify specific DNA sequences

Gel electrophoresis

Separates DNA or proteins by size using an electric field

DNA microarray

Chip containing thousands of probes to measure gene expression.

RNA‑seq

Sequencing‑based method to quantify all RNA transcripts.

Cluster analysis

Grouping genes or samples based on expression similarity.

Gene expression profiles

Patterns of gene activity across conditions or tissues.

Nuclear Magnetic Resonance & Mass Spectrometry

Analytical tools for identifying molecular structures and metabolites.

Why do we use high‑throughput technologies for tissue engineering?

To rapidly analyze thousands of genes, proteins, or metabolites at once, revealing how cells respond to scaffolds, signals, and environments.

What is the process of cell differentiation?

Cells activate lineage‑specific genes, produce new proteins, and adopt specialized structures and functions.

How do we sequence DNA?

Most commonly via chain‑termination (Sanger) or next‑generation sequencing (NGS). In Sanger, ddNTPs terminate DNA synthesis at specific bases. In NGS, millions of fragments are sequenced in parallel. Both require DNA polymerase, primers, and detection of nucleotide incorporation.

How is DNA sequencing becoming more efficient?

Through next‑generation sequencing (NGS) and third‑generation technologies (e.g., PacBio, Oxford Nanopore). Efficiency gains come from: parallelization (millions of reactions at once), reduced reagent volumes, automation, and longer read lengths—drastically lowering cost and time per genome.

How is DNA sequencing related to Polymerase Chain Reaction (PCR)?

PCR amplifies specific DNA regions to generate enough material for sequencing. Many sequencing library preparations include a PCR step to add adapters and increase DNA quantity. Conversely, sequencing can validate PCR products (e.g., checking for correct gene edits).

What methodologies are used in transcriptomics?

Main methods: DNA microarrays (hybridization‑based, probes on a chip) and RNA‑seq (next‑generation sequencing of cDNA). Other techniques include qPCR (low‑throughput) and single‑cell RNA‑seq (scRNA‑seq) for cellular heterogeneity.

How is transcriptomics used in tissue engineering

To monitor gene expression changes during differentiation, compare engineered tissue to native tissue, identify off‑target effects of scaffolds or growth factors, and characterize cell phenotype. Helps optimize culture conditions and verify tissue maturity before implantation.

What methodologies are used in proteomics?

Mass spectrometry (MS) – especially LC‑MS/MS – is the core method. Others include two‑dimensional gel electrophoresis (2D‑PAGE), protein microarrays, and NMR. MS identifies and quantifies thousands of proteins from cell lysates or secreted matrix.

How is proteomics used in tissue engineering?

To quantify extracellular matrix (ECM) proteins, signaling proteins, and contractile machinery in engineered tissues. Reveals post‑translational modifications, protein turnover, and cell‑matrix interactions. Helps assess whether a tissue has the correct protein composition and function.

Primary Culture

Cells taken directly from living tissue (e.g., biopsy) and placed in culture conditions for the first time. They retain many original characteristics but have limited lifespan. Used in tissue engineering to closely mimic native cell behavior.

Fluorescence‑activated cell sorter (FACS)

A technique that uses fluorescent labels and electric charges to rapidly separate cells based on size, granularity, and protein markers. In tissue engineering, _______ isolates specific cell populations (e.g., stem cells) or removes dead cells before seeding scaffolds.

Organ culture

A culture method where whole organs or tissue fragments are maintained with preserved three‑dimensional architecture and cell‑cell interactions. Used to study organ development, drug responses, or as a bridge to engineering whole organs.

Primary explant culture

A type of primary culture where small pieces of tissue (explants) are placed directly onto a culture surface. Cells migrate out of the explant and grow. Common for establishing primary cells from skin, cartilage, or cornea.

Cell culture

The process of growing and maintaining cells outside their natural environment, under controlled conditions (temperature, pH, nutrients). The foundation of tissue engineering for expanding cells before seeding onto scaffolds.

Passage or subculture

The transfer of a small number of cells from an existing culture to a new vessel with fresh medium. Used to expand cell numbers.

Cell Line

A population of cells that can be propagated repeatedly in culture, often derived from a single cell type. Can be finite (dies after a set number of divisions) or continuous (immortalized). Examples: HeLa, NIH/3T3.

Cell senescence

The irreversible arrest of cell division that occurs after a finite number of doublings.

Hayflick limit

The finite number of times a normal somatic cell population can divide before entering senescence. In humans, about 40–60 divisions. Important for planning primary cell expansion.

Telomerase

An enzyme that adds repetitive DNA sequences (telomeres) to the ends of chromosomes, preventing shortening during replication. Most somatic cells lack _________; its activation can immortalize cells for long‑term tissue engineering studies.

Immortalized cell line

A cell line that has escaped normal senescence and can divide indefinitely, often through genetic mutations or telomerase expression. Useful for reproducible experiments but may behave differently from primary cells.

Cell culture shock

Stress responses (e.g., growth arrest, apoptosis, altered gene expression) caused by sudden changes in culture conditions—temperature, pH, osmolality, or medium composition. Minimized by gradual adaptation and proper handling.

Confluence

The percentage of the culture vessel surface covered by adherent cells. 100% _________ means the surface is completely covered. Contact inhibition stops further growth. Used to time passaging or differentiation induction.

Oncogenes

Genes that have the potential to cause cancer when mutated or overexpressed. In tissue engineering, introduction of certain ______ (e.g., myc, ras) can immortalize cells but also risks transformation; used cautiously.

Transformed cell lines

Cell lines that have acquired genetic changes leading to unlimited growth, loss of contact inhibition, and often tumorigenicity. Can arise spontaneously or via viruses/chemicals. Useful for high‑throughput assays but not for implantation.

Genotypic drift

Accumulation of genetic changes (mutations, rearrangements) in cultured cells over time due to replication errors or selection. Leads to differences from the original tissue. Monitored by short tandem repeat (STR) profiling.

Phenotypic drift

Gradual changes in cell morphology, marker expression, or function during prolonged culture without genetic changes (epigenetic or selective). Example: chondrocytes losing cartilage‑matrix production after many passages.

Media

The nutrient solution that supports cell growth, containing amino acids, vitamins, salts, glucose, and often serum or defined additives. Tissue engineering uses specialized media to promote differentiation (e.g., osteogenic, chondrogenic media).

Blood serum

The liquid fraction of blood after clotting, rich in growth factors, hormones, and attachment proteins. Commonly fetal bovine serum (FBS) is added to media (5–20%) to promote cell proliferation. In tissue engineering, serum‑free defined media are increasingly used for clinical safety.

Substrate

The liquid fraction of blood after clotting, rich in growth factors, hormones, and attachment proteins. Commonly fetal bovine serum (FBS) is added to media (5–20%) to promote cell proliferation. In tissue engineering, serum‑free defined media are increasingly used for clinical safety.

Teratoma

A tumor containing tissues from all three germ layers (ectoderm, mesoderm, endoderm). In tissue engineering, _______ formation is an in vivo test for pluripotency of stem cells (e.g., iPSCs) before clinical use.

Cryopreservation

The process of cooling and storing cells or tissues at very low temperatures (typically −80°C or −196°C in liquid nitrogen) to halt biological activity. Essential for long‑term banking of primary cells, stem cells, and engineered constructs.