Introduction to Lab Techniques Measurements and Calibration

System Internationale (SI) system  Seven basic properties 1. Length 2. Mass 3. Time 4. Electrical current 5. Thermodynamic temperature 6. Luminous intensity 7. Amount of substance. SI units are related in a systematic fashion by using PREFIXES

 Biological scientists measure both physical properties (e.g.. mass, temperature) AND chemical/biological properties (e.g. enzyme activity).  National Institute for Standards and Technology (NIST) maintains >1300 standard reference materials whose compositions are determined as exactly as possible using modern methods.  Methods must also be “standardized” amongst individuals e.g. US PharmacopoeiaUS Pharmacopoeia

Calibration  …” to adjust a measuring system so that the values it gives are in accordance with external standard(s)”….  Manufacturer calibrates the instrument and end user or service technician must also recalibrate repeatedly….why?  Error in calibration is called tolerance. E.g. 500g +/- 1.2mg  Maintaining instruments “in calibration” is critical in the laboratory.

Calibration (standard) curves.  Relates to chemical and biological assays  Shows graphically the relationship between the response of the instrument and the amount of reference standard present.  Constructing a standard curve 1. Prepare a series of known concentrations of the material of interest. 2. Measure the response of the instrument to each standard 3. Plot the response of the instrument on the Y axis and the concentration on the X axis 4. Standard curve is used to determine the concentration of material in the unknown samples.

Accuracy and Precision  it is possible for a series of measurements to be precise but not accurate…………..why?  Definitions:  Precision: consistency of a series of measurements or tests (repeatability and reproducibility).  Accuracy: how close a measurement is to the true or accepted value (assess using a standard).  Understanding the difference is key in interpreting data and information!

 Accuracy can be expressed mathematically in 2 ways: 1. ABSOLUTE ERROR = (True value) – (average measured value). 2. % ERROR = (true value) – (measured value) X 100% (true value) If the results lie outside of an accepted range then it is necessary to look for problems……….troubleshooting!! The fun part!!

Significant Figures  A significant figure is the digit within a number that is a reliable indicator of value.  Table illustrates the “rules”.

Measurement of Weight  Weight is the force of gravity on an object.  Mass is the amount of matter in an object expressed in units of grams (independent of gravity)  Instruments: mechanical or electronic balances.  Errors: vibrations, drafts, static, temp. changes.  Calibration: ASTM protocol and a series of NIST weight standards; every day for GLP.

1. Mechanical Balances  Typically have one or more beams  Object is balanced against standards of known weight (NIST).  Inexpensive and easy to use.  No electrical signal.

2. Electronic Balances  Use electromagnetic force to counterbalance the sample.  Magnitude of electronic signal is related to sample’s weight.  Easy to use, interface with computer  automatic documentation.

Measurement of Volume  “….amount of space a substance occupies”  Instruments: Beakers, Graduated cylinders, volumetric flasks, Erlenmeyer flasks, pipettes, burettes, micropipettes.  Errors: technique, liquid viscosity, temperature  Calibration: capacity marks and graduation marks determined using ASTM standards and procedures. - TC (to contain) or TD (to deliver)

Using Micropipettors 1. Technique is most important factor affecting accuracy. 2. Physical and chemical properties of liquid will affect volumes delivered. 3. Environmental conditions! 4. Condition of micropipettor. Major concern is cross-contamination creation of aerosols, use filtered tips and good technique.

Measurement of Temperature  “……….measure of the average energy of the randomly moving molecules in a substance”.  Instruments: Liquid expansion devices,Bimetallic expansion devices, Change-of-state indicators, Metallic resistance devices.  Errors: depends on type of thermometer  Calibration: ASTM standards followed; check against a “certified reference thermometer”; reference thermometer calibration should be 10x more precise than the working thermometer.

Comparison of Fahrenheit, Celsius and Kelvin temperature scales. Absolute zero-460 o F-272 o C0K Freezing point of water 32 o F0oC0oC273K Average room temp. 68 o F20 o C293K Normal Human temp o F37 o C310K Boiling point of water 212 o F100 o C373K