Standard Operating Procedure for Glass Washing and Detergent Residues Test SOP Number: QC-03-07 Date Revised: 04-16-14


US Environmental Protection Agency
Office of Pesticide Programs
Office of Pesticide Programs
Mic ro bio lo gy La bo ra to ry
Environmental Science Center, Ft. Meade, MD
Standard Operating Procedure for
Glass Washing and Detergent Residues Test
SOP Number: QC-03-07
Date Revised: 04-16-14

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SOP No. QC-03-07
Date Revised 04-16-14
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SOP Number
QC-03-07
Title
Glass Washing and Detergent Residues Test
Scope
This protocol describes procedures for washing laboratory glassware
and for shipping glassware to a contract laboratory for analysis.
Detergents used in washing glassware may leave residues which are
bacteriostatic. If residues are present, glassware may require
additional rinsing to remove them (see section 15).
Application
To verify that detergent residue is not present in laboratory
glassware when cleaned using the dishwashers.

Approval Date
SOP Developer:

Print Name:
SOP Reviewer

Print Name:
Quality Assurance Unit

Print Name:
Branch Chief

Print Name:


Date SOP issued:

Controlled copy
number:

Date SOP withdrawn:

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SOP No. QC-03-07
Date Revised 04-16-14
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TABLE OF CONTENTS
Contents	Page Number
1.
DEFINITIONS
3
2.
HEALTH AND SAFETY
3
3.
PERSONNEL QUALIFICATIONS AND TRAINING
3
4.
INSTRUMENT CALIBRATION
3
5.
SAMPLE HANDLING AND STORAGE
3
6.
QUALITY CONTROL
3
7.
INTERFERENCES
3
8. NON- C ONF ORMIN G DATA
3
9.
DATA MANAGEMENT
3
10.
CAUTIONS
3
11.
SPECIAL APPARATUS AND MATERIALS
4
12.
PROCEDURE AND ANALYSIS
4
13.
DATA ANALYSIS/CALCULATIONS
9
14.
FORMS AND DATA SHEETS
9
15.
REFERENCES
10

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SOP No. QC-03-07
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1. Definitions
Abbreviations/definitions are provided in the text.
2. Health and Safety
Follow procedures specified in SOP MB-01, Laboratory Biosafety.
The Study Director and/or lead analyst should consult the Material
Safety Data Sheet for specific hazards associated with products.
3. Personnel Qualifications
and Training
Refer to SOP ADM-04, OPP Microbiology Laboratory Training.
4. Instrument Calibration
Not Applicable.
5. Sample Handling and
Storage
1.	Use and store detergents according to manufacturer's
instructions.
2.	Refer to section 10 for cautions associated with handling of Petri
dishes.
6. Quality Control
For quality control purposes, the required information is documented
on the appropriate form(s) (see section 14).
7. Interferences
1.	Inspect all glassware prior to use. Discard items with chips and
etched surfaces.
2.	Ensure dishwashers are working properly prior to commencing
the assay (e.g., scheduled preventive maintenance, if available).
3.	Petri plates should be shipped within 36 hours after completion
of sterilization.
8. Non-conforming Data
1.	Management of non-conforming data will be consistent with
SOP ADM-07, Non-Conformance Reports.
2.	Any deviation from the protocol will be documented. If the
regular wash procedure (Group A plates) is found to be
inadequate for removal of inhibitory detergent residues, then the
wash procedure will be adjusted and the detergent residue test
repeated.
9. Data Management
1.	Data will be archived consistent with SOP ADM-03, Records
and Archives.
2.	The plates are analyzed by a contract laboratory qualified to
perform detergent residue testing (for example; QC Laboratories,
Southhampton, PA). The reports of inhibitory residue analysis
received from the contract laboratory and completed forms must
be placed in the Quality Assurance of Glass Washing and
Detergent Residues Test Book.
10. Cautions
1. Ensure plates are properly packaged in a cardboard box with
suitable packing material prior to shipping to contract laboratory.

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SOP No. QC-03-07
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2. Laboratory personnel should run full dishwasher loads whenever
possible, and consider wash programs that conserve water and
produce glassware or labware appropriate for its intended use.
11. Special Apparatus and
Materials
1.	Miele Thermal Disinfector/Laboratory Glassware Washer Model
G7783 serial number 16/18344823 located in room B206
(Glassware and Media Preparation Room) of the OPP
Microbiology Laboratory at the ESC, Ft. Meade, MD.
2.	Lancerl600 UP Laboratory Glassware Washer serial number
9G050714 located in room B206 (Glassware and Media
Preparation Room) of the OPP Microbiology Laboratory at the
ESC, Ft. Meade, MD.
3.	Powder Detergent for Miele dishwasher
4.	Liquid Detergent for Lancer dishwasher
5.	Alconox Powdered Precision Cleaner
6.	Glass Petri Dishes (20 x 100 mm)
7.	Cardboard box and suitable packing material
8.	Disposable plastic Petri dishes provided by the contract
laboratory
12. Procedure and Analysis
Perform the detergent residues test annually or when a new lot or
different type of detergent is used. The test bacteria used is
Enterobacter aerogenes.
1.	All rinses use recycled rinse water from the washer reservoir.
2.	Glassware is washed in the Miele Thermal
Disinfector/Laboratory Glassware Washer and the Lancerl600
UP Laboratory Glassware Washer.
3.	Use the detergent as specified by the manufacturer.
4.	Hand-washed items will be analyzed when hand washing is
actively being utilized.
5.	Wash each group (Group A, Group B and Group C) separately in
each dishwasher.
6.	The procedure includes four groups of six Petri dishes each.
Group A, Group B and Group C contain Petri dishes washed by
dishwashers and Group D contains sterile plastic Petri dishes as
a reference point.
7.	Preparation numbers are assigned to each group of Petri dishes

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SOP No. QC-03-07
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according to SOP QC-15, Media Prep and Sterilization Run
Numbers.
8.	Group A (six glass Petri dishes) are washed and rinsed by the
regular procedure used in the laboratory for the particular
dishwasher.
9.	Group B (six glass Petri dishes) are washed by the regular
procedure used in the laboratory followed by twelve additional
rinses.
10.	Group C (six glass Petri dishes) are washed with the detergent at
the dilution normally used in the laboratory but the Petri dishes
are not run through any rinse cycles.
11.	Group D (six sterile, plastic Petri dishes) are provided by the
testing laboratory as a reference point.
12.1 Washing Procedure
for Miele Thermal
Dis infe ctor/Laborato ry
Glassware Washer
(Model G7783)
Group A (six glass Petri dishes):
a.	Place the Petri dishes in the dishwasher facing down and spaced
evenly so the water will run out of the dish. Place three Petri
dishes in the lower compartment and three Petri dishes in the
upper compartment.
b.	Fill the detergent compartment on the door with the amount
suggested by the manufacturer of powder detergent (2 scoops)
and close the detergent cover. In addition, place one scoop of
detergent directly on the washer door and a second one on the
base of the dishwasher for the Pre-wash cycle. A total of 4
scoops are used. Record the amount of detergent used on the
media prep sheet.
c.	Press the button for Program E, the Universal wash program,
and then press the Start button (diamond with vertical bar
symbol). This program includes a pre-wash and heated main
wash (85°C), two tap water rinses and two DI water rinses, one
unheated and one heated (70°C). This will constitute the normal
treatment that all machine-washed laboratory glassware receives
in this dishwasher.
Group B (six glass Petri dishes):
a. Place the Petri dishes in the dishwasher facing down and spaced
evenly so the water will run out of the dish. Place three Petri
dishes in the lower compartment and three Petri dishes in the
upper compartment.

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b.	Fill the detergent compartment on the door with the amount
suggested by the manufacturer of powder detergent (2 scoops)
and close the detergent cover. In addition, place one scoop of
detergent directly on the washer door and a second one on the
base of the dishwasher for the Pre-wash cycle. A total of four
scoops are used. Record the amount of detergent used on the
media prep sheet.
c.	Press the button for Program E, the Universal wash program,
and then press the Start button (diamond with vertical bar
symbol). This program includes a pre-wash and heated main
wash (85°C), two tap water rinses and two DI water rinses, one
unheated and one heated (70°C). This will constitute the normal
treatment that all machine-washed laboratory glassware receives
in this dishwasher.
d.	To achieve the additional twelve rinses for Group B, repeat the
following steps twelve times:
e.	Turn the dishwasher off at the end of the cycle.
f While pressing the T1 and T2 buttons, turn the dishwasher back
on.
g.	Select Program D. Press the T2 button so that the temperature is
set to 60°C. It may be necessary to press the T2 button a few
times to set the temperature to 60°C.
h.	Continuously press the start button until "38" appears in the
display. Note: As the display climbs from 1 to 38, you will hear
various components in the machine cycle on and off This is
normal.
i.	Washer will automatically stop at the end of the cycle
(approximately 12-15 minutes). Repeat steps 12.1 i-1 a total of
twelve times.
Group C (six glass Petri dishes):
a.	Place the Petri dishes in the dishwasher facing down and spaced
evenly so the water will run out of the dish. Place three Petri
dishes in the lower compartment and three Petri dishes in the
upper compartment.
b.	Fill the detergent compartment on the door with the amount
suggested by the manufacturer of powder detergent (2 scoops)
and close the detergent cover. In addition, place one scoop of
detergent directly on the washer door and a second one on the

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SOP No. QC-03-07
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base of the dishwasher for the Pre-wash cycle. A total of four
scoops are used. Record the amount of detergent used on the
media prep sheet.
c.	Press the button for Program E, the Universal wash program,
and then press the Start button (diamond with vertical bar
symbol).
d.	The washer must be stopped after the main wash cycle but
before the first tap water rinse, approximately 20 minutes after
starting the Universal cycle when the digital display reads 20
minutes, thus eliminating the two tap water rinses and two DI
water rinses.
12.2 Washing Procedure
for Lancer 1600 UP
Laboratory Glassware
Dishwasher
Liquid detergent is dispensed automatically through a metering
pump at the rate of approximately 5 mL/sec in this dishwasher. The
water consumption estimated per operation (according to the baskets
used) is 30 L. The metering pump during normal wash is set to run
for approximately 75 seconds to deliver the appropriate volume of
detergent.
Group A (six glass Petri dishes):
a.	Place the Petri dishes in the dishwasher facing down and spaced
evenly so the water will run out of the dish. Place three Petri
dishes in the lower compartment and three Petri dishes in the
upper compartment.
b.	Enter 10 on the keypad to select Cycle 10 and press the Start
button. Cycle 10 is the designated standard laboratory wash
program. This program includes a pre-wash and heated main
wash (85°C), two tap water rinses and three DI water rinses, two
unheated and one heated (70°C).
Group B (six glass Petri dishes):
a.	Place the Petri dishes in the dishwasher facing down and spaced
evenly so the water will run out of the dish. Place three Petri
dishes in the lower compartment and three Petri dishes in the
upper compartment.
b.	Enter 10 on the keypad to select Cycle 10 and press the Start
button. Cycle 10 is the designated standard laboratory wash
program. This program includes a pre-wash and heated main
wash (85°C), two tap water rinses and three DI water rinses, two
unheated and one heated (70°C).
c.	To achieve the additional twelve rinses for Group B, run Cycle

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30 four times. Cycle 30 consists of 3 cold demineralized rinses.
Enter 30 on the keypad to select Cycle 30 and press the Start
button.
Group C (six glass Petri dishes):
a.	Place the Petri dishes in the dishwasher facing down and spaced
evenly so the water will run out of the dish. Place three Petri
dishes in the lower compartment and three Petri dishes in the
upper compartment.
b.	Enter 20 on the keypad to select Cycle 20 and press the Start
button. Cycle 20 includes a pre-wash and heated main wash
(85°C) without the two tap water rinses and three DI water
rinses.
12.3 Hard Washing
Procedure (when
needed)
a.	Group A: wash six glass Petri dishes in Alconox detergent at the
manufacturers' prescribed dilution (1%), rinsed with tap water,
and then two times with DI water.
b.	Group B: wash six glass Petri dishes in Alconox detergent at the
manufacturers' prescribed dilution (1%), rinsed with tap water,
and then two times with DI water. To achieve the additional
twelve rinses for Group B, rinse the Petri dishes twelve
additional times with DI water (use DI water as it comes from
the DI faucet not the DI water from the Barnstead unit).
c.	Group C: wash six glass Petri dishes in Alconox detergent at the
manufacturers' prescribed dilution (1%) omitting the tap water or
DI rinses.
12.4 Petri Plate Preparation
and Sterilization
a.	Wrap each group of dishes in brown paper obtained from room
A140 (Region 3).
b.	Record group designation and dishwasher type (Miele A, Miele
B, Miele C, Lancer A, Lancer B, Lancer C, Handwash A,
Handwash B or Handwash C) on outside of paper.
c.	Sterilize all plates for each Group in the usual manner (gravity
cycle for 25 min).
d.	Record the sterilization run number on the appropriate
Laboratory Detergent Residue Test Form (see section 14).

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SOP No. QC-03-07
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12.5 Shipping
a.	After sterilization, place the wrapped plates in a cardboard box
with suitable packing material to prevent the plates from
breaking during shipment.
b.	Obtain a chain of custody form (COCform) from the contract
laboratory's website (for example QC laboratories). Fill the
form with all the necessary information. The original COC form
is placed inside the box with plates; a copy is retained for
internal records.
c.	Use a shipping company that is authorized by the agency.
Obtain shipping company's airbill from MLB authorized
personnel and fill with all necessary information. Shipping
should be conducted overnight or next day to ensure plates arrive
at the contract laboratory within 36 hours after sterilization (see
section 7).
d.	Notify the ESC mailroom personnel of the package to be
shipped.
e.	Ship package to contract laboratory, for example: QC
Laboratories, 1205 Industrial Blvd., Southampton PA 18966-
0514 and the phone number as 215-355-3900.
f Notify contract laboratory that package is being shipped for
Detergent Residue Analysis (for example, QC Laboratory phone
number 215-355-3900).
13. Data Analysis/
Calculations
1.	When the report of inhibitory residue is received from the
contract laboratory, record the testing results legibly and in
indelible ink under the "Calculations and Conclusions" column
(see section 14).
2.	Differences in averaged counts on plates in Groups A through D
should be less than 15% if there are no toxic or inhibitory
effects.
3.	Differences in averaged counts of less than 15% between
Groups A and B and greater than 15% between Groups A and C
indicate that the cleaning detergent has inhibitory properties that
are eliminated during routine washing.
4.	Differences between B and D greater than 15% indicate an
inhibitory residue.
14. Forms and Data Sheets
Test Sheets. Test sheets are stored separately from the SOP under
the following file names:

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SOP No. QC-03-07
Date Revised 04-16-14
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Laboratory Detergent Residue Test Form QC-03-07 F1 docx
for Machine Washed Items ~
Laboratory Detergent Residue Test Form „ „ __ ,
for Hand Washed Items QC-03-07_F2.doc*
15. References
1.	Bordner, R. FL, J. A. Winter and P. V. Scarpino. eds. 1978.
Microbiological Methods for Monitoring the Environment,
Water and Wastes. EPA-600/8-78-017, Environmental
Monitoring and Support Laboratory, U.S. Environmental
Protection Agency, Cincinnati, Ohio.
2.	Eaton, A. D., Clesceri, L. S., Rice, E. W., Greenberg, A. E. and
Franson, M. A. H. eds. 2005. Standard Methods for the
Examination of Water and Wastewater, 21st Edition. American
Public Health Association, Washington, DC.

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