Uploaded by Dr John Grifson

PIPETTE anuba

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GLASSWARES
AND
PIPETTES
CONTENTS
• GLASS-TYPES
• GLASSWARES
• PIPETTES
GLASS
 Glass is a complex silicate compound
 Properties depends on silicate anion as well as its
cation content
 Addition of metal ions
BOROSILICATE GLASS
Ingredients
•Silica
Ingredients
•Boron
Silica oxide
•Sodium
oxide
Boron oxide
•Aluminium
Sodium oxide oxide
• Aluminium oxide
Percent
80.6
12.6
4.15
2.2
FEATURES
 High thermal resistance
 Low Alkali content
 Free from Zinc group elements and heavy metals
 Resists heat, corrosion and thermal shock
 Low coefficient of expansion
FEATURES
Thermal durability of borosilicate glass:
Strain point
510 °C
Annealing point
555 °C
Softening point
820 °C
USES
Advantages:
Can be heated upto 600 °C
Brand names: Pyrex, Kimax
Most lab equipment
Beakers,flasks,pipettes
COREX-ALUMINIUM SILICATE
GLASS
• Resist clouding due to alkali ,staining
• Harder
• For temperature more than250 c
• 30000 PSI-6 times stronger than borosilicate
• Thermometers,graduated cylinders,centrifuge
tubes
HIGH SILICA GLASS
Made by removing all elements except silica
from borosilicate glass
High silica content – 96%
Comparable with fused quartz in thermal
endurance, chemical stability and electrical
characteristics
Radiation resistant
Good optical qualities
USES
Used for optical reflectors, cuvettes , high
temperature thermometers
VYCOR
 Used in applications involving high
temperatures, drastic heat shock and
extreme chemical treatment
 Resistant to attack by all acids except
hydrofluoric acid
Uses:
 Ashing and ignition techniques
BORON FREE GLASS
 Less than 0.2% boron
 High resistance to alkali
 Thermal resistance less
Uses: strong alkali reactions
LOW ACTINIC GLASSWARE
 Impart red or amber colour to glass
• Uses:
 Provides highly protective laboratory
glassware for handling materials sensitive
to light in the 300 – 500 nm range
• Eg: Bilirubin, Carotene, Vitamin A
FLINT GLASS
Soda lime glass – mixture of silicon,
calcium, sodium oxides
Lowest cost
Poor resistance to high temperatures and
sudden changes of temperature
• Uses:
Bottles, disposable pipette
DISADVANTAGES
• Breakable
• May leak alkali in solution
COLOURED AND OPAL
GLASSES
 Made by adding small amounts of metallic
oxides to the glass
Uses:
 Filters and light bulbs
OPTICAL GLASS
 Made of lead, sodalime and borosilicate
 High optical purity
Uses:
 Prisms, lens, mirror
COATED GLASS
 Has thin metallic oxide permanently
firebonded to the surface of glass
 Conducts electricity
• Uses:
 Heat shield to protect against infra red light
 Electrostatic shield to carry off charge
GLASS CERAMICS
High thermal resistance, chemical stability, and
corrosion resistance
Useful for making hot plates, heat exchangers
RADIATION ABSORBING GLASS
Made of soda lime and lead
Useful in preventing the transmission of high
energy rays (X-rays and γ rays)
ASTM DESIGNATIONS
• Class A glassware:
Highest level of accuracy
Borosilicate is used
• Class B glassware:
Same basic design as class A
Sodalime is used
Volumetric tolerances for class B – twice the
specified range allowed for class A
LABORATARY GLASSWARE
• VOLUMETRIC
• GRADUTED
CYLINDERS
• VOLUMETRIC FLASKS
• BURETS
• PIPETTES
• NON VOLUMETRIC
• FUNNELS
• CUVETTES
• TEST TUBES
• DESSICATORS
• SYRINGES
• REAGENT BOTTLE
• STIRRING ROD
• PETRI DISH
• WATCH GLASS,COVER SLIPS
VOLUMETRIC FLASKS
 Flat-bottomed flask ,round lower portion
 Long thin neck
 Etched calibration line
 To prepare reagents
 single graduation line
 Class A
• For accurate volumetric measurements
• Pipette to be used in adding final drops
TOLERANCE TABLE
Capacity (mL)
25
50
100
250
500
1000
Tolerance for
class A(mL)
0.03
0.05
0.08
0.12
0.20
0.30
Tolerance for
class B(mL)
0.06
0.10
0.16
0.24
0.40
0.60
ERLENMEYER FLASKS
• Wide bottom,evolves into smaller short
neck
• Chemicaly inert and heat resistant
• Preparing solutions and titrations
• Not accurate
ROUND SHAPED FLASK
• Round Shaped Flat Bottomed Flask:
Often used to evaporate a sample to dryness
Round Shaped Round Bottomed flask:
Used in distillation
Should not be heated
Not to be used for reagent storage
BURETTES
Long graduated glass tubes with a stop cock at one end
Used to deliver accurately, known amounts of liquid into
the container
BURETTE
Standard burettes vary in size from 25 to 100 mL
Stop cock – all glass or all Teflon at one end
Micro processor controlled automatic burette with
accuracy 0.1%
Digital display - reading upto 0.001mL
USES
• For titrations
• Known volume is dispensed
BEAKERS
 Wide mouthed, straight sided, cylindrical vessels
Volumes vary from 5mL to several liters
 The Griffin beaker - flat bottom, straight sides,
and an opening as wide as the flat base, with a
small spout in the lip.
• Uses:
General mixing
Non volumetric liquid reagents preparation
GRADUATED CYLINDERS
Narrow long ,cylindrical tubes
Circular or octagonal base
Measure volume of liquids
Not as accurate as volumetric flask
Size vary from 10-2000ml
FUNNELS
• Filtering funnels:
58 – 60 ° angled funnels with either short or long thin
stems
• Powder funnels:
For transferring solids
Wide mouthed stem
• Separatory funnels:
 Pear shaped or cylindrical
 Used for manual liquid – liquid extractions
CUVETTES
CUVETTES
 Special kinds of test tube with straight sides
 May be rectangular in shape
 Holds solutions intended for photometric readings
 Width determines the length of the light path through the
sample
 Have two transparent sides opposite one another
 Costly
 Should not get scratched
DESSICATORS
Used to dry solids or liquids
Dessicant is placed in the bottom
A shelf is placed above the dessicant
in which the material is placed
Wide, flat ground glass lip on the top
CENTRIFUGE TUBES
SYRINGES
Used for accurate volumetric work
Injection of small volume of sample,
• liquid or gas for chromatography
REAGENT BOTTLES
PIPETTES
PIPETTE
• Used to transfer a known volume of liquid
from one container to another
• creates a partial vacuum above the liquid
holding chamber and selectively releases this
vacuum to draw up and dispense liquid
TYPES OF PIPETTES
TC - TD PIPETTES
• Difference is the amount of liquid needed to wet
the interior surface of the ware
• The amount of any residual liquid left in the pipet
tip.
• Most manufacturers stamp TC or TD near the top of
the pipet to alert the user as to the type of pipet.
• TC pipet holds or contains particular volume but
does not dispense that exact volume,
• TD pipet will dispense the volume indicated.
TC PIPETTE
 Contains particular volume
 Doesn’t dispense the same volume
TD PIPETTE
• Dispenses the volume indicated
BASED ON DRAINAGE
CHARACTERISTICS
• Self draining pipette
• Blow out pipette
BASED ON GRADUATION
MEASURING / GRADUATED PIPETTE
Mohr pipette
Serological pipette
MEASURING PIPETTE
• Piece of glass tubing that is drawn out to a
tip and graduated along its length
• MOHR pipette-calibrated between two
marks
• SEROLOGICAL pipette-graduated to the tip
MOHR PIPETTE
SEROLOGICAL PIPETTE
Smaller orifice
Larger orifice
Needs more controlled delivery Drains faster
Blow out to deliver
entire volume
DIFFERENCE
MOHR PIPETTE
SEROLOGICAL PIPETTE
MEASURING PIPETTE
• Room for error
• Only used to measure reagents
TRANSFER PIPETTES
• Designed to transfer known volume
• Volumetric pipettes
• Ostwald –folin pipettes
VOLUMETRIC PIPETTES
• To deliver accurately a fixed volume of aqueous
solution.
• Precision decreases with decrease in size
• Cylindrical bulb joined at 2 ends to narrower glass
tubes
• Caliberation on upper suction tube
• Delivery tube is tapered
• To prevent uncontrolled delivery and incomplete
drainage
OSTWALD- FOLIN PIPETTE
• Similar to volumetric pipette
• The bulb is closer to the delivery tip
• For accurate measurement of viscous fluid, blood,serum.
• Etched mark to indicate blow out pipette
• Slow drainage
MICROPIPETTE
• TO measure microliter volumes
• Volume coating inner walls can cause error
• TC pipettes
• Rinsed with the final solution after delivering contents
into the diluent
• 1-1000ul
FIXED VOLUME AND VARIABLE
VOLUME PIPETTE
MULTI CHANNEL PIPETTE
TYPES-MECHANISM
• Air-displacement pipettes
relies on a piston for suction creation to
draw the sample into a disposable tip
• Positive-displacement pipettes
Piston moves like in a hypodermic syringe
POSITIVE DISPLACEMENT PIPETE
• Direct contact with the liquid
• Air cushion in pipette tip
• Displaces the liquid
USES
• Viscous liquid
• Volatile liquid
• Hot or cold
PIPETTING TECHNIQUE
• Set the volume
• Attach the tip
• Depress the plunger to first stop
• Immerse the tip into the liquid and draw liuid by
releasing piston
• Dispense the liqud-push plunger top
• Withdraw pipette before releasing the piston
• Remove tip by ejecting
DONT’S WHEN PIPETTING
• Never use pipette without pipette tip
• Do not use pipette beyond volume limits
• Never repeatedly jam tip into a pipette if it doesn’t fit
• Don’t push past first stop while aspirating set volume
• Release push button in a controlled manner
• Always make sure tip is below liquid level while aspirating
• Never place pipette horizontally without expelling liquid
PHOTOMETRIC CALIBRATION
• Method I :
 An aliquot of diluent is pipetted out
 The change in concentration will reflect the volume of the
pipette
• Method II :
Comparing absorbances of dilutions of
potassium dichromate using Class A
volumetric glassware versus equivalent
dilutions made with the pipetting device
GRAVIMETRIC CALIBERATION
• MATERIALS :
• Pipette
• 10 to 20 pipette tips
• Balance
• Weighing vessel
• Type I water
• Thermometer
METHOD
• Record weight of vessel
• Note temperature and pressure
• Place small volume of water in container
and cover
CALIBERATION
• Set the balance to zero
• Dispense specified amount by pipette to be
tested
• Record result
• Repeat 9 more times after changing tips
CALCULATION
CALCULATE
• VOLUME OF WATER
 MEAN WEIGHT
•
•
Z-DENSITY OF WATER AT GIVEN
TEMPERATURE AND PRESSURE
CALCULATION
• VOLUME ( ml or µl ) = WEIGHT( mg or µg ) x Z
ACCURACY AND PRECISION
• Accuracy is the proximity of measurement
results to the true value;
• precision is the degree to
which repeated (or reproducible)
measurements under unchanged
conditions show the same results.
……………..
ACCURACY & PRECISION
ACCURACY TOLERANCES
• CLASS A
• Accuracies in mL of Manual pipettes
Type of
pipette
1.0 mL
NBS
standard
5.0 mL
10.0 mL
25.0 mL
0.01
0.02
0.025
Class A
Volumetric
0.006
0.01
0.02
0.03
Mohr
0.01
0.02
0.03
0.10
Serological
0.01
0.02
0.03
0.10
SEMIAUTOMATIC AND
AUTOMATIC PIPETTES
• Programable devices
• Dispensing alquots of liquid into multiple
wells
• Piston driven
ELECTRONIC PIPETTES
• Electronically dispenses predefined
volumes(0.5-200ul)
• Disposable plastic tips to contain
• Retains less in inner walls
• Avoids contamination
• Improves reliability
AUTOMATIC DISPENSING
APPARATUS
• Aspirates & dispenses preset volume of
two different liquids
• 2 motor driven syringes
• Microprocessor controlled
• Easily programmed and stored in memory
PERSONAL COMPUTER CONTROLLED
APPARATUS
ROBOTIC LIQUID HANDLING
WORK STATION
• For clinical,genomic,proteomic applications
• 96-384 well microtiter plates
• Single or multi probed
• 2 dimensional movement of probes
• 0.5-1000ul in serial dilutions 4-16 channels
ROBOTIC LIQUID HANDLING
WORKSTATION
ROBOTIC LIQUID HANDLING
WORKSTATION
BULBS
TIPS
Commonly available pipette volumes
Min. volume
(µL)
Max. volume
(µL)
P2
0.2
2
Orange
10
P10
1
10
Red
10
P20
2
20
Lemon
200
P100
20
100
Salmon
200
P200
50
200
Yellow
200
P1000
200
1000
Blue
1000
P5000
500/1000
5000
Purple
5000
P10000
1000
10000
Sky
10000
Name
Color on Gilson[clarification
needed]
tip size (µL)
PIPETTING TECHNIQUES
• Tip must be immersed in liquid to be
transferred
• Without touching the vessel wall
• Pipet is held upright, not at an angle
• Pipet bulb or similar device used to apply
slight suction
READING MENISCUS
NBS
• National bureau for standards-standard
specifications for glass volumetric
apparatus
Class A Borosilicate volumetric glassware
• Superior thermal and chemical resistance
• Better suited - glassware for storage of
solutions, e.g. flasks.
• Accuracy is retained over a longer working
lifetime than their soda-lime equivalents.
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