Unit 1 lab techniques
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| Describe how to carry out a serial dilution | 1ml of stock solution with 9ml of water. Then 1ml of diluted solution with 9ml of water and so on....
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| What substance controls pH | buffers
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| Describe two techniques that can be used to quantify the concentration of an unknown solution | 1.colorimetry 2. titration
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| What is another term for a colorimeter | spectrophotometer
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| What is a standard curve | measuring absorbance of known concentrations in a colorimeter to extrapolate unknown from graph
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| On what property are substances separated in centrifugation | density
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| What name is given to the substance at bottom and top during centrifugation | bottom - pellet top - supernatant
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| Small substances are found where after centrifugation | in supernatant
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| What technique separates proteins based on pH | Iso electric focusing
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| What is meant by the iso electric point of a protein | When it is OVERALL neutral and precipitates out of solution
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| What are the two types of gel electrophoresis by which proteins can be separated | 1. Denaturing 2. Native/non denaturing
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| Explain the process of denaturing gel electrophoresis | All proteins are denatured to give a negative charge and move to positive end dependent on size
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| Explain the process of native gel electrophoresis | Proteins remain in original conformation (no denaturing) and migrate through gel dependent on SIZE & CHARGE
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| What type of gel electrophoresis depends only on size | Denaturing (as all proteins are artifically made negative)
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| What type of gel electrophoresis depends on size and charge | Native (protein charge not affected)
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| Name a factor affecting protein migration apart from size or charge | size of pores in buffer
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| What are the types of microscopy | 1. bright field 2. Fluorescence
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| Which type of microscopy allows tiny structures such as individual proteins within the cell to be viewed | Fluorescence
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| Which type of microscopy is more limited in its resolution | Bright field
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| Which type of microscopy is more simple to use | Bright field
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| Which type of microscopy involves antibodies to tag objects | Fluorescence
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| Name an object that can be viewed under a bright field microscope | whole organisms, parts of organism or thin tissues
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| What does a haemocytometer estimate | Total cell count
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| What technique is able to distinguish between dead cells and viable cells | Vital Staining with trypan blue
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| Explain why only dead cells stain with trypan blue | Trypan blue toxic so actively pumped out by living (viable) cells
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| Explain how to estimate total cell count in a haemocytometer | Ensure all units are in CM. L x B X Depth = volume
Count cells that fit in grid
Proportion calculation to scale up for chosen volume which is always larger than volume calculated
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| What term is given to cells that can only divide a limited number of times (hayflick) limit | primary cell lines
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| What term is given to cells that can divide infinitely | Cancer cells
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| Why are growth regulators/hormones used in plant tissue | To enable explant to divide into callus AND to enable callus to differentiate into plantlet
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| Why do mammalian cells need foetal bovine serum | For cell proliferation
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| Why do mammalian cells need antibiotics | to kill bacteria which grow more readily than mammalian cells
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| What type of organisms are produced from inoculum | microbes
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| What type of organisms are produced from explants | plants
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| Describe the 5 key stages of mammalian cell sub culture | Use proteolytic enzymes to remove mammalian cells from tissue
Cells adhere to surface
Cells flatten
Cells divide to form a monolayer
Cells form confluent
Process repeats
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| What is a protoplast | plant cell without cell wall
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| How are different species of plants produced in a lab | 2 protoplasts fused together with electric shock & cell wall reformed
plant growth regulators then added to divide cells to form callus which differentiate into plantlet
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| Name a plant hormone/growth regulator | auxin or cytokinins
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| Name a substance a plant needs to grow except plant hormones | M & S salts, Macroelements NPK or Microelements Cu, Zn, vitamins, glucose
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| What two cells fuse to make a hybridomas | B lymphocyte with a myeloma cell
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| How are specific monoclonal antibodies produced in a lab | injecting mouse with specific antigen that produces antibodies. Removing spleen and extract B lymphocytes & fusing with myeloma cell to form hybridomas
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| What substance are the hybridomas placed in and subsequently diluted to ensure one cell per well | polyethylene glycol (PEG).
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