1 Workshop 1: A Practical Approach to Aeroallergen Identification Estelle Levetin, PhD How to Set Up a Sampling Station Learning Objectives and Disclosure Information Upon completion of this workshop, participants should be able to: Set up a sampling station to collect airborne pollen and fungal spore Recognize the most common types of pollen found in the atmosphere Recognize the most common types of fungal spores found in the atmosphere No conflicts to disclose Aerobiological Sampling Sampling plan or objective Choosing samplers: Rotorod, Burkard Spore Trap, Lanzoni, Allergenco One day head or 7-day head for Burkard or Lanzoni Location Preparing the samples Slide Analysis and Identification Data Analysis ACAAI Annual Meeting Nov. 7 - 11 2013, Baltimore
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A Practical Approach to Aeroallergen Identification
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Workshop 1: A Practical Approach to Aeroallergen Identification
Estelle Levetin, PhD
How to Set Up a Sampling Station
Learning Objectives and Disclosure Information Upon completion of this workshop, participants
should be able to: Set up a sampling station to collect airborne pollen and fungal
spore
Recognize the most common types of pollen found in the atmosphere
Recognize the most common types of fungal spores found in the atmosphere
Models most often used by allergists have retracting rods for intermittent operation
Standard is 10% sampling time
Head rotates at 2400 rpm
Leading edge of rod coated with grease
Pollen and spores impacted on greased surface
Efficient for pollen and spores >10 m
ACAAI Annual MeetingNov. 7 - 11 2013, Baltimore
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Rotorod Samplers
Older Model Rotorod Sampler
Rotorod Analysis
Collector rods placed in a special adapter for microscopic examination
Rods stained with Calberla’s pollen stain
Entire surface of each rod counted unless pollen/spore load very high (then a subset of the surface is analyzed)
Atmospheric concentrations determined
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Rotorod Calculations
C = N / VC is concentration, N is the total number of pollen or spores counted on both rods, V is the volume of air sampled by the rods
V = Rod area (m2) x D x x RPM x tRod area = width of rod (1.52 mm = 0.00152m) x length of the rod (23 mm = 0.023m) x 2 (both rods), D is the diameter of the Rotorod head (8.5 cm = 0.085m), RPM is 2400, t is minutes sampled per day
With a 5% sampling time (72 min) V = 3.226 m3
Concentration = N/3.226 m3
With a 10% sampling time (144 min) V = 6.452 m3
Concentration = N/6.452 m3
Hirst Spore Trap
Burkard Spore Trap
Lanzoni Spore Trap
Allergenco
Burkard and Lanzoni Spore Traps
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Allergenco – Samplair MK-3
Advantages of Burkard Spore Trap
High efficiency down to less than 5 m Allows for greater
accuracy for small fungal spores
Time discrimination Permits analysis for
diurnal rhythms
Permanent slides for future reference
Location
Roof of a building - ideal 3 to 6 stories above ground (30 to 60 ft)
Not close to overhanging vegetation
Air flow not obstructed by nearby buildings or other structural features
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Parapet around roof requires platform to elevate the orifice above the wall
Telescoping mast elevates sampler above local vegetation.
Burkard 7-day sampler head
Standard is the 7-day sampling head
Sampler drum mounted on 7-day clock
Drum moves by orifice at 2 mm per hr
Melenex tape mounted on drum and greased (Lubriseal, High Vacuum Grease, other)
Air is brought in at 10 l/min and impacts on greased Melenex tape
Drum changed each week
Seven Day Sampling Head
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Processing the 7-day drum
Melenex tape removed from drum Tape cut into seven 24 hour segments each 48 mm
long Segments mounted on microscope slides in 10%
gelvatol (polyvinyl alcohol) and dried Glycerin-jelly mounting medium added and a 50 mm
cover slip Mounting medium contains pollen stain - either basic
fuchsin or phenosafarin
Melenex tape on cutting board
One-day sampling head
Alternate head is the 24 hour head Standard glass microscope slide is greased
and placed on the head Alternatively Melenex tape can be fixed on the
slide and greased
Slide is changed daily, carrier realigned Mounting medium with stain and coverslip
are added
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24 hour sampling head
Outdoor air sample from Tulsa
Analysis
Microscopy - 400X for pollen; 1000X for fungal spores
Different methods of microscopic analysis are used to obtain Average daily concentration - Single longitudinal
traverse
Hourly or bihourly concentrations which can then be averaged to obtain a daily average - 12 transverse traverses
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The Single Longitudinal Traverse Method
The Twelve Transverse Traverse Method
Burkard Counting Methods
Comparison of methods
Single Longitudinal Traverse Quicker
Produces average daily concentration
Good for routine monitoring
3 or 4 longitudinal traverses can increase accuracy
12 Transverse Traverses Takes longer
Can determine diurnal rhythm of airborne allergens
All traverses can be averaged to determine average daily concentration
Conversion to Concentrations
Microscope counts are entered into a database such as Excel
Formulas added to convert counts into concentrations
Information needed Field diameter of objective lens - Variable Flow rate (10 liters/minute) and exposure time (normally 24
hrs) for a total volume of air sampled of 14.4 m3
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Calculating Concentrations for Single Longitudinal Traverse
C = Concentration - pollen grains/m3
N = number of pollen counted on traverse W = Width of entire sample - 14 mm F = field diameter of objective lens - 0.48 mm V = total volume of air sampled- 14.4 m3