Mol bio lab Flashcards
is concerned with the organizational process and the conditions under which laboratory studies are planned, performed, monitored, recorded, and reported.
Good laboratory (GLP)
Closeness of agreement between a measured quantity value and a true quantity value of a measurand
ACCURACY
Particular quantity intended to be measured
Example: vapor pressure of a given sample of water at 20°C.
Measurand
Also called “dead volume”, the air cushion is the volume of air located between the lower part of the pipette piston and the surface level of the sample.
AIR CUSHION
Measured portion of a homogeneous entity. A general term referring to multiple samples of any solution, mixture, etc.
ALIQUOT
An instrument for delivering predetermined volumes of liquid from a reservoir. The reservoir may be integrated into the instrument or connected externally.
DISPENSER
Rounded or approximate value of characterizing quantity of a measuring instrument or measuring system that provides guidance for its appropriate use.
Examples: a) 1 L as the value marked on a single-mark volumetric flask, b) 100 μL as the setting appearing on the volumeter of a pipette.
NOMINAL VALUE
Closeness of agreement between indications or measured quantity values obtained by replicate measurements on the same or similar objects under specified conditions.
PRECISION
An instrument for transferring a predetermined volume of liquid from one vessel to another. A pipetter is not connected to a reservoir
PIPETTE/ PIPETTER
The appropriate representative part of a liquid to be analyzed. The term “test sample “is used when necessary to avoid confusion with the statistical term “random sample from population”
SAMPLE
The portion of the sample that is retained in the instrument after sample delivery and that may affect subsequent samples.
NOTE: Carryover from a positive displacement pipette is less than from an air- displacement pipette.
SAMPLE CARRYOVER
Total volume and temperature range, as well as ambient conditions, for which instrument performance is specified.
Note: do not select volumes outside recommended limits.
WORKING RANGE
Measurement precision under reproducibility conditions of measurement.
Reproducibility (of Results of Measurements)
Types of Pipettes:
Air displacement
Positive displacement
pipetting is highly accurate for standard pipetting applications.
Air displacement
pipetting is based on direct contact of the piston with the liquid.
Positive displacement
Recommended for aqueous solutions, such as buffers, diluted acids, or alkalis, this technique is commonly used when pipetting and mixing a sample or reagent into another liquid.
Forward Technique
This technique is commonly used with air displacement pipettes, and is recommended for precisely pipetting small volumes. Reverse pipetting avoids the risk of sample splash, foaming, or bubble formation.
Reverse Technique
This technique is intended for repeat dispensing of the same volume, and is ideal for adding reagents into tubes or wells of microplates.
Repetitive Pipetting Technique:
is a multi-purpose tip for many laboratory applications with a variety of performance requirements that range from high accuracy to reagent dispensing with greater tolerance.
A standard tip
Cross-contamination from pipetting is a constant concern within a lab. Two options exist within the Thermo Scientific Portfolio: Filter Tips and Barrier Tips
Sterile filter and barrier tips
utilizes pore size and treacherous path technologies to help control liquid from accidentally splashing the inside of a pipette.
A filter tip
utilizes a self-sealing technology to prevent liquid from passing and accidentally splash the inside of a pipette.
An ART Barrier tip
are recommended for low volume applications in genetic studies, forensics and PCR or anytime contamination is a concern.
Filter and Barrier tips
Utilizing polymer technology makes the inner surface of the pipette tip more hydrophobic, resulting in a reduction in sample loss due to adhesion. Benefits include improved sample delivery and conservation of expensive reagents.
Low retention pipette tips
are designed for unique pipetting applications to save time, reduce contamination, and increase accuracy, precision, and productivity.
Specialty tips
feature a distal end orifice that is nearly 70 percent larger than that of a standard pipette tip. These tips provide the added flexibility required for handling difficult-to-pipette samples.
They are designed for researchers working with macromolecules like genomic DNA and are especially critical for transferring fragile cellular samples such as macrophages, hybridomas, and hepatocytes, as well as other viscous materials.
Wide orifice tips
allow you to access the bottom of test tubes, reagent bottles, flasks, and other vessels without touching the shaft of the pipette against the side of the tube. This adds a layer of security to protect samples, and virtually eliminates the chance of carryover contamination.
The longer tip length allows you to reach the bottom of long or narrow vessels that standard tips cannot reach.
Extended length tips
Loading acrylamide or agarose gels with standard pipette tips can be a time- consuming process. Use the round gel loading tips for agarose gels and specialized Ultra Round and Ultra Flat gel tips for your polyacrylamide gels to speed up the loading process.
8. Solvent-safe carbon filtered pipette tips are the best solution for handling the pipetting rigors of Combinatorial Chemistry. These specialized tips keep strong acids, bases, and aggressive organic solvents from causing pipette failures and critical inaccuracies.
Gel loading tips
are the best solution for handling the pipetting rigors of Combinatorial Chemistry. These specialized tips keep strong acids, bases, and aggressive organic solvents from causing pipette failures and critical inaccuracies.
Solvent-safe carbon filtered pipette
Pipette maintenance is done by either the manufacturer or the end-user. For maintenance, always check the manual and documentations that come with the pipette.
Pipette Basic Maintenance
is a procedure used to isolate DNA from
the nucleus of cells.
DNA extraction
To obtain DNA in a relatively purified form which can be used for further investigations, i.e. PCR, sequencing
Non-ionic detergent , Salt, Buffer, EDTA designed to lyse outer cell membrane, but will not break down nuclear membrane.
Cell Lysis Buffer
is a chelating agent of divalent cations such as Mg2+. Mg2+is a cofactor for DNase nucleases. If the Mg2+is bound up
by EDTA, nucleases are inactivated.
EDTA (Ethylenediaminetetraacetic disodium salt)
it is usual to remove most of the protein by digesting with proteolytic enzymes such as proteinase K, which are active against a broad spectrum of native proteins, before extracting with organic solvents. Proteinase K is approximately 10 fold more active on denatured protein. Proteins can be denatured by SDS or by heat.
Proteinase K
remove proteinaceous material.
Phenol/Chloroform
is often used to store DNA and RNA. EDTA in TE chelates Mg2+ and other divalent metals ions necessary for most causes of DNA and RNA degradation, suppressing these processes. Tris is a buffering agent to keep the solution at a defined pH.
TE Buffer - Tris-EDTA Buffer: TE buffer
which help in precipitation of DNA.
Ethanol or isopropanol
This buffer will lyse
cell membrane, nuclei are intact, pellet nuclei.
1.Cell Lysis Buffer
The stages of the method are
Lyse
Bind
Wash
Elute
The cells of a sample are broken open with a lysis procedure
Lyse
A buffer solution is then added to the sample along with ethanol or isopropanol. This forms the binding solution. The binding solution is transferred to a spin column and the column is put in a centrifuge. The centrifuge forces the binding solution through a silica gel membrane that is inside the spin column. If the pH and salt concentration of the binding solution are optimal, the nucleic acid will bind to the silica gel membrane as the solution passes through.
Bind
The flow-through is removed and a wash buffer is added to
the column. The column is put in a centrifuge again, forcing the wash buffer through the membrane. This removes any remaining impurities from the membrane, leaving only the nucleic acid bound to the silica gel.
Wash
The wash buffer is removed and an elution buffer (or simply water) is added to the column. The column is put in a centrifuge again, forcing the elution buffer through the membrane. The elution buffer removes the nucleic acid from the membrane and the nucleic acid is collected from the bottom of the column.
Elute
