The Real Deal Midterm 1
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291 Terms
What is magnification?
The enlargement of an object's apparent size.
What is resolution?
The ability to distinguish two objects that are close together as separate objects.
What is the diffraction limit of light microscopy?
Approximately 200 nanometers or 0.2 micrometers.
What is phase contrast microscopy used for?
Visualizing live
What is differential interference contrast (DIC) microscopy used for?
Visualizing live
What is fluorescence microscopy?
A technique that uses fluorophores to visualize specific molecules or structures in cells.
What is a fluorophore?
A molecule that absorbs light at one wavelength and emits light at a longer wavelength.
Why do cells often need to be labeled for microscopy?
Cells are typically colorless
What is DAPI?
A fluorescent dye that stains DNA and is commonly used to visualize nuclei.
What are the three major ways to label cells for fluorescence microscopy?
Fluorescent dyes
What is immunofluorescence?
A technique that uses fluorophore-conjugated antibodies to visualize specific proteins in cells.
What is an antigen in immunofluorescence?
The specific protein that an antibody recognizes and binds.
What is a primary antibody?
An antibody that specifically recognizes and binds the protein of interest.
What is a secondary antibody?
An antibody that recognizes and binds the primary antibody and is conjugated to a fluorophore.
Why use a fluorophore-conjugated secondary antibody?
It allows the location of the primary antibody and its target protein to be visualized.
What does immunofluorescence tell you about a protein?
Where the protein is located within a cell.
Why are cells fixed before typical immunofluorescence?
Fixation preserves cellular structure and helps keep proteins in place during labeling.
Why are cells permeabilized during immunofluorescence?
To allow antibodies to enter the cell and access intracellular proteins.
What is the purpose of blocking during immunofluorescence?
To minimize nonspecific antibody binding and background fluorescence.
What does colocalization of two fluorescently labeled proteins indicate?
The proteins are found in the same location or cellular region.
What is a fluorophore-tagged protein?
A protein of interest genetically fused to a fluorescent protein such as GFP.
How are fluorophore-tagged proteins produced?
A transgene encoding the protein of interest fused to a fluorescent protein is introduced and expressed in cells.
What is GFP?
Green fluorescent protein
What does imaging a GFP-tagged protein allow researchers to observe?
The protein's expression
What is a fluorescent fusion protein?
A protein produced from a gene encoding a protein of interest joined to a fluorescent protein.
What is a transcriptional reporter?
A construct in which a promoter of interest drives expression of a fluorescent reporter
What is the difference between a fluorescent protein fusion and a transcriptional reporter?
A fusion reports the protein's location and behavior; a transcriptional reporter indicates activity of a particular promoter.
What does scanning electron microscopy (SEM) visualize?
The topography and composition of a specimen's surface.
What does transmission electron microscopy (TEM) visualize?
Internal structures of thinly sliced
Why can electron microscopy reveal smaller structures than light microscopy?
Electrons have shorter wavelengths than visible light and therefore greater resolving power.
What type of specimen preparation is required for TEM?
A fixed specimen that is thinly sliced so electrons can pass through it.
What type of specimen preparation is required for SEM?
A fixed specimen prepared for imaging its surface.
What is cell fractionation?
The process of breaking open cells and separating their components into different fractions.
What is differential centrifugation?
A technique that separates cell components based on differences in size and density using progressively faster centrifugation.
During differential centrifugation
which components generally pellet first?
During differential centrifugation
which components require faster centrifugation to pellet?
What is a cell homogenate?
The mixture produced when cells are broken open
What is a pellet in differential centrifugation?
The material that collects at the bottom of the centrifuge tube.
What is a supernatant in differential centrifugation?
The liquid remaining above the pellet after centrifugation.
How can researchers identify which organelle is enriched in a cell fraction?
They can use antibodies against proteins known to be associated with particular organelles.
How can immunoblotting be used after differential centrifugation?
Separate fractions are analyzed with antibodies to determine which fraction contains a protein of interest.
What does a band on an immunoblot indicate?
The presence of the protein recognized by the antibody at a particular molecular weight.
Why include a whole-cell lysate control in an immunoblot experiment?
To confirm that the protein was present in the original cells before fractionation.
What is the function of the nucleus?
It stores the cell's genome and is the site of DNA replication and transcription.
What is the nuclear envelope?
A double membrane surrounding the nucleus.
What are nuclear pores?
Protein complexes in the nuclear envelope that regulate transport between the nucleus and cytosol.
What is the function of mitochondria?
They harvest energy from food molecules and generate ATP through cellular respiration.
What are the two membranes of a mitochondrion?
The outer membrane and the inner membrane.
What is the mitochondrial intermembrane space?
The compartment between the outer and inner mitochondrial membranes.
What is the mitochondrial matrix?
The compartment enclosed by the inner mitochondrial membrane.
Where is the mitochondrial electron transport chain located?
In the inner mitochondrial membrane.
Where is mitochondrial ATP synthase located?
In the inner mitochondrial membrane.
Why is the inner mitochondrial membrane highly folded?
Its folds increase the surface area available for energy-harvesting proteins and ATP synthesis.
What is the function of chloroplasts?
They harvest light energy to produce ATP and NADPH and use these molecules to help convert CO2 into organic compounds.
What are the internal membrane compartments of chloroplasts?
Thylakoid membranes
Where are the electron transport chain and ATP synthase located in chloroplasts?
In the thylakoid membrane.
What is the stroma of a chloroplast?
The fluid-filled compartment surrounding the thylakoids.
What is the thylakoid lumen?
The internal space enclosed by the thylakoid membrane.
What is the endosymbiotic theory?
The theory that mitochondria and chloroplasts evolved from bacteria that were engulfed by ancestral eukaryotic cells.
What evidence supports the endosymbiotic origin of mitochondria and chloroplasts?
They have their own DNA
What is the function of lysosomes?
They contain digestive enzymes that break down and recycle cellular materials.
What is the lysosomal lumen?
The internal compartment of a lysosome
What is the function of peroxisomes?
They contain enzymes involved in oxidative reactions
What is a major difference between lysosomes and peroxisomes?
Lysosomes digest and recycle cellular materials; peroxisomes carry out oxidative reactions.
What is the endoplasmic reticulum (ER)?
An extensive membrane network involved in protein and lipid synthesis and processing.
What is the rough ER?
The portion of the ER studded with ribosomes that synthesize proteins entering the secretory pathway or membranes.
What is the smooth ER?
The portion of the ER lacking ribosomes and involved in lipid synthesis and other specialized functions.
What is the function of the Golgi apparatus?
It modifies
How is the Golgi apparatus structurally organized?
As a stack of flattened membrane-enclosed discs.
What is the cytosol?
The aqueous fluid portion of the cytoplasm outside membrane-enclosed organelles.
What is the cytoplasm?
The contents of the cell inside the plasma membrane
What are the three major cytoskeletal filament systems in animal cells?
Actin filaments
What are actin filaments?
Thin cytoskeletal filaments involved in cell shape
What are microtubules?
Hollow cytoskeletal tubes involved in intracellular transport
What are intermediate filaments?
Strong
What does it mean that membrane lipids are amphipathic?
They have both a hydrophilic region and a hydrophobic region.
Why do amphipathic membrane lipids spontaneously form bilayers in water?
Their hydrophilic heads interact with water while their hydrophobic tails avoid water and pack together.
What are the four types of membrane lipids covered in the lecture slides?
Glycerophospholipids
What is a glycerophospholipid?
A membrane lipid with a glycerol backbone
What is a sphingolipid?
A membrane lipid built on a sphingosine backbone rather than glycerol
What is a glycolipid?
A membrane lipid with one or more sugar molecules attached to its head group.
What is a major function of glycolipid sugar groups?
They contribute to cell recognition and signaling.
Where do glycolipid sugar head groups face?
Toward the extracellular environment.
What is cholesterol's role in the membrane?
It buffers membrane fluidity by limiting excessive movement at high temperatures and preventing tight packing at low temperatures.
How does fatty acid chain length affect membrane fluidity?
Shorter fatty acid tails increase fluidity; longer tails decrease fluidity.
How does fatty acid saturation affect membrane fluidity?
Unsaturated tails increase fluidity; saturated tails decrease fluidity.
Why do unsaturated fatty acid tails increase membrane fluidity?
Their double bonds create kinks that prevent lipids from packing tightly together.
Why do saturated fatty acid tails decrease membrane fluidity?
Their straight tails pack closely together
How do longer fatty acid tails affect interactions between lipids?
They increase interactions between neighboring tails
What happens to membrane fluidity when temperature increases?
Lipid movement increases and the membrane becomes more fluid.
What happens to membrane fluidity when temperature decreases?
Lipid movement becomes restricted and the membrane becomes less fluid.
What is the transition temperature (Tm) of a membrane?
The temperature at which the membrane transitions between a more rigid state and a more fluid state.
How does cholesterol affect membrane fluidity at high temperatures?
It restrains phospholipid movement and prevents the membrane from becoming excessively fluid.
How does cholesterol affect membrane fluidity at low temperatures?
It disrupts tight phospholipid packing and helps prevent the membrane from becoming too rigid.
What does it mean that cholesterol buffers membrane fluidity?
It reduces extreme changes in fluidity as temperature changes.
What is the fluid mosaic model?
The model describing membranes as flexible lipid bilayers containing proteins that can move laterally.
What does it mean that a membrane is a two-dimensional fluid?
Lipids and many proteins can move laterally within the plane of the bilayer.
What are integral membrane proteins?
Proteins that penetrate the lipid bilayer
What are transmembrane proteins?
Integral membrane proteins that span the entire lipid bilayer.
What are peripheral membrane proteins?
Proteins associated with the membrane surface through noncovalent interactions without entering the bilayer.