calibrationofglassware-120408004156-phpapp02.ppt
- 1. Analytical Chemistry BS (HONS) Chemistry University Of Gujrat Hafiz Muhammad Athar Rizwan Roll # 09050607-008 1
- 2. Presentation Topic Calibration of Glassware 2
- 3. Basics Review and Calibration of Volumetric Glassware There are three types of containers used in lab to contain or deliver liquids: volumetric, ordinary, and disposable glassware. Volumetric glassware are containers that have been calibrated at a specific temperature to deliver or contain very precise amounts of liquid. Examples of volumetric glassware that we will use include burettes, pipettes, and volumetric flasks. Ordinary glassware has less precise volume calibrations and are used whenever the volumes do not have to be measured as accurately. Examples include beakers, Erlenmeyer flasks, and graduated cylinders. Disposable glassware (or plastic ware) is used to transfer or hold liquids temporarily and may not contain any volume markings. Examples include medicine droppers and disposable pipettes. 3
- 4. Volumetric pipette or flask, that is, you will experimentally determine what volume a pipette or flask really delivers. You will also calibrate a beaker or Erlenmeyer flask and a 50 ml burette. You will use these calibrated values throughout the remainder of the quarter. Care of glassware: Glassware should be washed with a mild soap solution, rinsed with tap water, and then rinsed with distilled water. If beads of water stick to the inside walls, it be should be cleaned again. If glassware is to be dried, allow it to drain or use paper towels. Never sling glassware to remove water. Never tap glassware against the side of the sink. A burette or pipette should be rinsed with a small amount of the solution to be used to remove any water droplets from the glass walls. 4
- 5. Burette: Burette must be read to the hundredths of a milliliter. Note that the liquid level markings begin at the top or open end. This is because the burette is designed to deliver liquids. All volumes are measured as the difference between an initial and final reading. Pipette: The volumetric pipette is designed to deliver a single, fixed volume of liquid at a specific temperature. A graduated pipette is used in a similar fashion to the burette to deliver specific amounts of liquid. Remember to touch off the drop of liquid hanging outside the tip into the transferred liquid (it is part of the delivered volume) but do not blow out any liquid that remains inside the tip of the pipette. The pipette has been calibrated to contain this last drop of liquid. 5
- 6. Principles of Measuring the Volume of Liquids • Overview – Volume • The amount of space a substance occupies • Liter – Basic unit of volume • Measuring devices – Depends of amount to measure and accuracy – Larger volumes » Glass and plastic vessels » Graduated cylinders » Volumetric flasks – Pipettes » Preferred for volumes in 1-25ml range – Micropipette devices » Less than 1ml 6
- 7. Basic principles of Glassware Calibration • Calibration up to manufacturer – Capacity marks/graduation • Lines marked on volume measuring devices that indicate volume – Lines the way to make the measurements – Determined by American Society for Testing and Materials (ATSM) • Meniscus – Must be considered – A curve formed by the surface of liquids confined in narrow spaces (measuring devices) » Lowest point of the meniscus used as point of reference for calibration » Hold at eye level to be entirely accurate 7
- 8. Basic principles of Glassware Calibration • Liquid measuring calibrated to either – To contain (TC) • Will contain the specified amount when filled to the capacity mark – Will not deliver that amount – Adhesion to sides – To deliver (TD) • Marked slightly differently so that it does deliver the specified amount, assuming the liquid is water at 20oC and it is poured using specific techniques – Volumetric flask » Hold specific volume » Essential for proper concentrations of buffers, etc » Not good for “delivering” proper volume » Use of TD device should be used 8
- 9. Basic principles of Glassware Calibration • Small amount error in calibration inevitable – Tolerance • How much error is allowed in the calibration of a volume measuring item • Most accurately calibrated glassware termed volumetric – Has narrowest tolerance allowed • Two effects of temperature to consider in high accuracy measurements – Expand or contract with changes in temperature • Affects accuracy – Volume of the solution changes as the temperature changes • Devices calibrated at 20oC with water 9
- 10. Glass and Plastic Labware used to Measure Volume • Beakers and Erlenmeyer flasks – Hold liquids – Not to measure • Calibrated with tolerance of ± 5% – 100mL line may be anywhere form 95-105mL • Graduated cylinders – Cylindrical vessels calibrated with sufficient accuracy for most volume measurements • ± 0.6mL tolerance • Usually calibrated TD • Can be obtained with different set of graduations, depending on need • Not designed for mixing or storing – Unstable/easily knocked over 10
- 11. Glass and Plastic Labware used to Measure Volume • Burettes – Long graduated tubes with a stopcock at one end that are used to dispense known volumes accurately • Volumetric Flasks – Vessels used to measure specific volumes where more accuracy is required than is attainable from a graduated cylinder • Calibrated either TC OR TD a single volume – Several disadvantages • Relatively expensive • Calibrated for only one volume • Used only in situations where high accuracy volume measurements are required 11
- 12. Glass and Plastic Labware used to Measure Volume • Volumetric Flasks – Proper use • Choose the proper type of flask for the application – TC or TD – Class A or Class B – Serialized or not • Be sure the flask is completely clean before use • Read the meniscus with eyes even with liquid surface • If calibrated TD, then pour as follows: – Incline the flask to pour the liquid; avoid splashing on the walls as much as possible – When the main drainage stream has ceased, the flask should be nearly vertical – Hold flask in vertical position for 30 sec or so and touch off the drop of water adhering to the top of the flask by touching it to the receiving vessel Never expose volumetric glassware to high temperatures because heat causes expansion and contraction that can alter it calibration 12
- 13. Pipettes-Pipettes and Pipette-Aids • Pipettes – Hollow tubes that allow liquids to be drawn into one end and are generally used to measure volumes in the 01.-25ml range • Glass or plastic • Disposable or multiple use • Pre-sterilize disposable with cotton plugged tops • Pipette-aids – Devices used to draw liquid into and expel it from pipettes • Mouth pipeting – Used to be the norm – Now banned by safety regulation 13
- 14. Pipettes-Measuring Pipettes • Measuring pipette – Calibrated with a series of graduation lines to allow the measurement of more than one volume • Serological pipettes – Usually calibrated so that the last drop in the tip needs to be “blown out” to deliver the full volume to deliver the full volume of the pipette. 14
- 15. Pipettes-Measuring Pipettes • Serological pipettes – To use: » Check that the pipette is calibrated to be “blown out” by looking for the bands at the top » Examine the pipette to be sure the tip is not cracked or chipped » Fill the pipette about 10mm above the capacity line desired and remove any water on the outside of the tip by a downward wipe with lint-free tissue » Place the tip in contact with a waste beaker and slowly lower the meniscus to the capacity line » Deliver the contents into the receiving vessel by placing the tip in contact with the wall of the vessel » When the liquid ceases to flow, “blow out” the remaining liquid in the tip with on firm “puff” with the tip in contact with the vessel wall. 15
- 16. Pipettes-Measuring Pipettes • Mohr pipettes – Calibrated “to deliver” but, unlike the serological pipettes, the liquid in the tip is mot part of the measurement and the pipette is not blown out. 16
- 17. Pipettes-Volumetric (Transfer) Pipettes • Volumetric (transfer) pipettes – Made of borosilicate grass and are calibrated TD a single volume when filled to their capacity line at 20oC. • Most accurately calibrated pipette • Can be serialized – Calibrated • “Delivery of the contents into the receiving vessel is made with tip in contact with the wall of the vessel and no after-drainage period is allowed. – Not “blown out” 17
- 18. Pipettes-Other Types • Pasteur pipettes – Used to transfer liquids from one place to another • Not volume-measuring devices – No calibration lines – Good for transferring liquids • Manual dispenser for reagent bottles – Devices placed in a reagent bottle with a tube that extends to the bottom of the bottle – Dispenser has a plunger that is depressed to deliver a set volume of liquid. 18
- 19. Micropipetting devices-positive displacement and air displacement micropipettors • Micropipettors – Devices commonly used to measure smaller volumes, in the 1- 1000µL range – Two types • Positive displacement – Include syringes and similar devices where the sample comes in contact with the plunger and walls of the pipetting instrument » Used for viscous and volatile samples • Air displacement micropipettors – Designed so that there is an air cushion between the pipette and the sample such that the sample only comes in contact with a disposable tip and does not touch the micropipettor itself » Disposable tips-reduce chance of cross contamination – Accurately measure volume of aqueous samples » Most common instruments used in biotech 19
- 20. Micropipetting devices-air displacement micropipettors • Obtaining accurate measurements – Procedure for operation • Plungers – Control used by operator for the uptake and expulsion of liquids » Different stop levels can be felt (fig 20.13, pg. 357) – Factor affecting accuracy • Operator’s technique • Physical/chemical properties of the liquids • Measurements affected by the environment in which they are made • Condition of the micropipettor 20
- 21. Glassware Laboratory glassware refers to a variety of equipment, traditionally made of glass, used for scientific experiments and other work in science, especially in chemistry and biology laboratories. Some of the equipment is now made of plastic for cost, ruggedness, and convenience reasons, but glass is still used for some applications because it is relatively inert, transparent, more heat-resistant than some plastics up to a point, and relatively easy to customize.0 ml. • Glassware can be divided into 2 groups – Non-volumetric glassware • Beaker • Flask – Volumetric Glassware • Volumetric Flask • Graduated Cylinder 21
- 22. Beaker • Used for transferring liquid to another container or to transfer a small amount of reagent for use in procedures. • Volume is not accurate, just an estimate. • Never place a reagent in another container without labeling the container first. 22
- 23. Erlenmeyer Flask • Features a conical base, a cylindrical neck and a flat bottom. • They are marked on the side (graduated) to indicate the approximate volume of their contents. • This is NOT used for ACCURATE measurement 23
- 24. Graduated Cylinder • For rapid measurement of liquid volume. • They are generally more accurate and precise for this purpose than flasks. • This is a semi-accurate liquid measuring vessels. 24
- 25. Reading the Volume • 10 mL has approx 6.62 mL • 100 mL 52.7 mL • 25 mL has 11.5 mL 25
- 26. Volumetric Flask • A volumetric flask is used to measure very precisely one specific volume of liquid (100 ml, 250 ml, etc., depending on which flask you use). • This flask is used to prepare a solution of known concentration. • To make up a solution, first dissolve the solid material completely, in less fluid than required to fill the flask to the mark. • After the solid is completely dissolved, very carefully fill the flask to the mL mark. • The top is then sealed and the flask is inverted several times to mix. 26
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