Agricultural Marketing Service

Science & Technology Program

Laboratory Approval and Testing Division

Laboratory Approval Service

Laboratory Approval Program – Mycotoxins

Doc. No: LAP-PR.07

Revision Date: 02/08/23 Page 1 of 29

1 Purpose

The purpose of this document is to outline the requirements for the Laboratory Approval 

Program for Analysis of Mycotoxins (LAP-Mycotoxins). The document describes the technical 

competency and quality management requirements a laboratory must demonstrate to be a USDAapproved laboratory. 

The Laboratory Approval Program (LAP) is administered by the Laboratory Approval Service 

(LAS) Branch. LAS is part of the Agricultural Marketing Service (AMS), Science and 

Technology (S&T) Program, Laboratory Approval and Testing Division (LATD). 

The LAS approves, or accredits, laboratories to perform testing services in support of domestic 

and international trade. At the request of industry, other Federal Agencies, or foreign 

governments, the LAS develops and administers programs to verify that the analysis of food and 

agricultural products meet country and customer-specific requirements and that the testing of 

products marketed is conducted by qualified and approved laboratories. 

2 Scope

This LAP is for a laboratory seeking to obtain and maintain its status as a USDA-approved 

laboratory for the analysis of mycotoxins in almonds, pistachios, and/or peanuts based on the 

stipulations of the final market destination: U.S. domestic, U.S. import, and export. This 

Program verifies technical and quality control competencies of a laboratory for the analysis of 

mycotoxins. All aspects of a laboratory’s quality management system (business processes 

relative to the scope of approval) are applicable and are critical for ensuring the defensibility of 

the analytical results produced under the LAP. 

2.1 Almonds for aflatoxins (B1, B2, G1, G2) for export to the European Union (EU) through the 

Pre-Export Certification program (PEC) of the Almond Board of California (ABC). 

2.2 Peanuts for aflatoxins (B1, B2, G1, G2) marketed domestically for human consumption, 

including imports, in accordance with 7 CFR 996. 

2.3 Peanuts for aflatoxins (B1, B2, G1, G2) for export to the EU. 

2.4 Pistachios for aflatoxins (B1, B2, G1, G2) for domestic and import markets in accordance 

with 7 CFR 983 and 7 CFR §999.600, respectively. 

2.5 Pistachios for aflatoxins (B1, B2, G1, G2) for export to the EU for the Administrative 

Committee for Pistachios’ (ACP) Pistachio Export Aflatoxin Reporting (PEAR) program. 

2.6 Pistachios for ochratoxin A (OTA) for export to the EU for the ACP PEAR program. 
Agricultural Marketing Service

Science & Technology Program

Laboratory Approval and Testing Division

Laboratory Approval Service

Laboratory Approval Program – Mycotoxins

Doc. No: LAP-PR.07

Revision Date: 02/08/23 Page 2 of 29

3 Table of Contents

1 Purpose......................................................................................................................................................... 1

2 Scope............................................................................................................................................................ 1

3 Table of Contents......................................................................................................................................... 2

4 Glossary of Terms ...................................................................................................................................... 2

5 References.................................................................................................................................................... 3

GENERAL REQUIREMENTS ...................................................................................................... 6

6 Laboratory Approval Program Administrative Policy ................................................................................. 6

7 Summary of General Program Requirements .............................................................................................. 6

8 Mandatory Quality Assurance Practices ...................................................................................................... 7

9 Method Selection ......................................................................................................................................... 7

10 Demonstration of Method Performance by Validation and Verification Evaluation ................................... 8

11 Analyst Competency and Proficiency Testing ............................................................................................. 9

TECHNICAL REQUIREMENTS (GENERAL) .......................................................................... 11

12 Performance Criteria ..................................................................................................................................11

13 Quality Controls.........................................................................................................................................11

14 Quality Measures .......................................................................................................................................12

15 Data Analysis.............................................................................................................................................14

16 Critical Equipment & Reagents..................................................................................................................15

17 Official Certificate of Analysis/Report ......................................................................................................16

TECHNICAL REQUIREMENTS (SCOPE SPECIFIC) .............................................................. 17

18 Almond – AFL – Export to EU..................................................................................................................17

19 Peanut – AFL – U.S. Domestic & U.S. Imports.........................................................................................19

20 Peanut – AFL – Export to EU....................................................................................................................21

21 Pistachio – AFL – U.S. Domestic & U.S. Imports.....................................................................................23

22 Pistachio – AFL – Export to EU ................................................................................................................25

23 Pistachio – OTA – Export to EU................................................................................................................27

24 Explanations for Technical Requirements ................................................................................................. 29

25 Revision History ........................................................................................................................................29

26 Review / Approvals....................................................................................................................................29

Table 1. Performance criteria for aflatoxins (B1, B2, G1, G2). ..................................................... 11

Table 2. Performance criteria for ochratoxin A. .......................................................................... 11

4 Glossary of Terms

ABC Almond Board of California

ACP Administrative Committee for Pistachios

AFL Aflatoxin

AMS Agricultural Marketing Service

AOAC Association of Official Analytical Collaboration International 

AOCS American Oil Chemists’ Society

APC American Peanut Council 

CFR Code of Federal Regulations 

CCV Continuing Calibration Verification

CV Coefficient of Variation
Agricultural Marketing Service

Science & Technology Program

Laboratory Approval and Testing Division

Laboratory Approval Service

Laboratory Approval Program – Mycotoxins

Doc. No: LAP-PR.07

Revision Date: 02/08/23 Page 3 of 29

EC European Commission 

EU European Union 

FAPAS Food Analysis Performance Assessment Scheme

FDA Food and Drug Administration 

FLD Fluorescence Detector

HPLC High Performance Liquid Chromatography 

IAC Immunoaffinity Column 

ICV Independent Calibration Verification 

ISO/IEC International Organization for Standardization/International Electrotechnical 

Commission

LAP Laboratory Approval Program 

LAP-Mycotoxins LAP for Analysis of Mycotoxins 

LAS Laboratory Approval Service 

LATD Laboratory Approval and Testing Division 

LC Liquid Chromatography (e.g., HPLC, UPLC) 

LOQ Limit of Quantitation

MOU Memorandum of Understanding 

OTA Ochratoxin A 

PEAR Pistachio Export Aflatoxin Reporting Program 

PEC Pre-Export Certification

PHRED Photochemical Reactor Enhanced Detection

PM Program Manager

PT Proficiency Testing 

S&T Science & Technology Program 

TLC Thin Layer Chromatography 

UPLC Ultra (High) Performance Liquid Chromatography 

USDA United States Department of Agriculture

VCM Vertical Cutter Mill

5 References

5.1 The following are referenced in this document. Dated references apply to the edition cited

and undated references apply to the latest edition published (including any amendments). 

5.2 Laboratory Approval Program: 

a) LAP-PR.05, Laboratory Approval Program – General Policies and Procedures.

b) LAP-PR.06, Laboratory Approval Program – Fees.

c) USDA AMS LATD LAS Website, https://www.ams.usda.gov/services/lab-testing/labapproval.
Agricultural Marketing Service

Science & Technology Program

Laboratory Approval and Testing Division

Laboratory Approval Service

Laboratory Approval Program – Mycotoxins

Doc. No: LAP-PR.07

Revision Date: 02/08/23 Page 4 of 29

5.3 Quality Assurance Standards: 

a) ISO/IEC 17025:2017. General requirements for the competence of testing and 

calibration laboratories.

b) USDA AMS Laboratory Standards of Practice. 

5.4 USDA Requirements: 

a) 7 CFR Part 981 – Almonds grown in California. 

b) 7 CFR Part 983 – Pistachios grown in California, Arizona, and New Mexico. 

c) 7 CFR Part 996 – Minimum quality and handling standards for domestic and imported 

peanuts marketed in the United States. 

d) 7 CFR §999.600 – Specialty Crops; Import Regulations. Regulation governing the 

importation of pistachios. 

e) Milled Peanuts, Inspection Instructions. USDA MRP AMS SCP SCID. December 

2020. 

f) Pistachio Nuts In the Shell Shipping Point and Market Inspection Instructions. USDA 

AMS, February 2005. 

g) Pistachio Nuts In the Shell Shipping Point and Market Inspection Instructions, Patch # 

045, Testing Samples For Aflatoxin. USDA AMS, April 15, 2019. 

h) Pistachio Nuts In the Shell Shipping Point and Market Inspection Instructions, Patch 

#066, Aflatoxin Sampling and Testing for Imported Raw and Blanched Pistachios. 

USDA AMS, April 28, 2021. 

5.5 FDA Requirements:

a) MOU 225-19-031. Memorandum of Understanding between AMS and FDA in 

inspecting, sampling, and testing peanuts, Brazil nuts, and pistachio nuts for aflatoxins. 

b) FDA Compliance Policy Guides, Section 570.375 Aflatoxin in Peanuts and Peanut 

Products. 

c) FDA Compliance Policy Guides, Section 570.500 Pistachio Nuts - Aflatoxin 

Adulteration. 

d) FDA Compliance Program Guidance Manual, 7307.001, Chapter 07 – Molecular 

Biology and Natural Toxins. Mycotoxins in domestic and imported foods FY 15/16. 

e) Guidelines for the Validation of Chemical Methods in Food, Feed, Cosmetics, and 

Veterinary Products, 3rd Edition. US FDA. FDA Foods Program, October 2019. 

5.6 Industry Requirements: 

a) Administrative Committee for Pistachios: Handler’s Guide. 

b) Almond Board of California, Pre-Export Checks (PEC) Program Manual. 
Agricultural Marketing Service

Science & Technology Program

Laboratory Approval and Testing Division

Laboratory Approval Service

Laboratory Approval Program – Mycotoxins

Doc. No: LAP-PR.07

Revision Date: 02/08/23 Page 5 of 29

c) Pistachio Export Aflatoxin Reporting (PEAR) Program. 

5.7 Methodology: 

a) AOAC International, Official Methods 977.16, Sampling for Aflatoxins - Preparation 

for Sample Procedure. 

b) AOAC International, Official Methods 991.31, Aflatoxins in Corn, Raw Peanuts, and 

Peanut Butter - Immunoaffinity Column (Aflatest) Method. 

c) AOAC International, Official Methods 998.03, Aflatoxins in Peanuts - Alternative BF 

Method. 

d) AOAC International, Official Methods 999.07, Aflatoxins B1 and Total Aflatoxins in 

Peanut Butter, Pistachio Paste, Fig Paste, and Paprika Powder – Immunoaffinity 

Column Liquid Chromatography with Post-Column Derivatization. 

e) AOAC International, Official Methods 2005.08, Aflatoxins in Corn, Raw Peanuts, and 

Peanut Butter - Liquid Chromatography with Post-Column Photochemical 

Derivatization.

f) Francis Jr., O. J. 1979. Sample Preparation of Some Shelled Tree Nuts and Peanuts in a 

Vertical Cutter-Mixer for Mycotoxin Analysis. JAOAC. 62(5):1182-5. 

5.8 Country Specific Regulations: 

a) EC 401/2006. Commission Regulation (EC) No. 401/2006 of 23 February 2006 laying 

down the methods of sampling and analysis for the official control of the levels of 

mycotoxins in foodstuffs. 

b) EC 1881/2006. Commission Regulation (EC) No. 1881/2006 of 19 December 2006 

setting maximum levels for certain contaminants in foodstuffs. 

c) EU 2015/949. Commission Implementing Regulation (EU) No. 2015/949 of 19 June 

2015 approving the pre-export checks carried out on certain food by certain third 

countries as regards the presence of certain mycotoxins. 

5.9 Additional Resources: 

a) Eurachem Guides.

b) Good Laboratory and Clinical Practices, Techniques for the Quality Assurance 

Professional, edited by P.A. Carson and N.J. Dent, 1990.
Agricultural Marketing Service

Science & Technology Program

Laboratory Approval and Testing Division

Laboratory Approval Service

Laboratory Approval Program – Mycotoxins

Doc. No: LAP-PR.07

Revision Date: 02/08/23 Page 6 of 29

GENERAL REQUIREMENTS 

6 Laboratory Approval Program Administrative Policy

6.1 A laboratory seeking admission to the LAP must fulfill the requirements and follow the 

process described in the program procedure, LAP-PR.05, Laboratory Approval Program - 

General Policies and Procedures. This procedure describes the process for application, 

assessment audits, acceptance, maintaining program status, suspension, withdrawal, 

dismissal, and appeals. 

6.2 This program is administered on an annual, calendar year, basis. 

6.3 The program procedure, LAP-PR.06, Laboratory Approval Program – Fees, explains the 

fees for services.

6.4 The administrative procedures above are available on the LAS website, or by contacting 

the LAS Mycotoxin Program Manager (PM) for the current version of the procedure. 

7 Summary of General Program Requirements

7.1 The laboratory must comply with all requirements set forth in this document to be 

compliant with the LAP. For a laboratory to maintain good standing, each year it must: 

a) meet all program requirements relevant to the scope of approval; 

b) comply with mandatory laboratory practices based on the ISO/IEC 17025 standard (See 

§8); 

c) use test method(s) approved by AMS. (See §9); 

d) validate analytical and sample preparation methods prior to use and ensure the 

validation data package is available for review upon request (See §10); 

e) evaluate analyst competency and maintain satisfactory status (See §11); 

f) participate in proficiency testing (PT) programs and maintain satisfactory status (See 

§11); 

g) submit sieve testing results to PM (See §14.3.h)); 

h) communicate regularly with the PM to share vital information regarding the laboratory 

and make all information relevant to the LAP available to PM upon request (See 

§5.2.a)); 

i) notify the PM within 30 days of significant changes relevant to the laboratory’s 

approval status and/or ability to meet the Program’s requirements including but not 

limited to: legal, organizational, or ownership status; laboratory policies, procedures, 

and resources; change in key managerial personnel, contact persons, and signatories; 
Agricultural Marketing Service

Science & Technology Program

Laboratory Approval and Testing Division

Laboratory Approval Service

Laboratory Approval Program – Mycotoxins

Doc. No: LAP-PR.07

Revision Date: 02/08/23 Page 7 of 29

and verification of adequate method performance after a change in location, equipment, 

facilities, and working environment. It is at the discretion of LAS whether an onsite 

visit/audit is required to evaluate any significant change that a laboratory undergoes 

(See §5.2.a)); 

j) be available for a biennial (every other year) re-assessment audit, have actual sample(s)

ready to demonstrate its competency of performing the test method, and comply with 

requests for documents and records before and during the audit (See §5.2.a)); 

k) respond to each nonconformance found with a record of an investigation, root cause 

analysis, and correction or, if warranted due to time, a corrective action plan, within 30 

calendar days of being reported (See §5.2.a)); 

l) resolve each nonconformance in a timely manner, whether identified by LAS auditor, 

another organization, or internally (See §5.2.a)); 

m) pay all program fees by the due date on the billing invoice (See §5.2.b)). 

8 Mandatory Quality Assurance Practices

8.1 Implement quality assurance and quality control procedures to ensure a validated and 

qualified analysis, prove competence, and ensure defensible data. This Program does not 

require ISO 17025 accreditation; however, it is a common practice for testing laboratories 

to become ISO 17025 accredited. 

8.2 LAS uses the ISO 17025 standard to evaluate all laboratory quality systems regardless of 

accreditation status. A nonconformance identified during a LAS assessment audit may be 

cited to the ISO 17025 standard (See §5.2.a)). 

8.3 Maintain records for at least three years (See §5.2.a)). 

9 Method Selection

9.1 Use analytical testing methods approved by AMS. 

9.2 Use method fit for purpose based upon the needs of the customer and consistent with 

specified requirements.

9.3 Use methods as validated; for the purposes of this Program, consider methods, with 

collaborative studies, specifically those published by USDA, FDA, and AOCS, as validated 

methods (see §5.7).

9.4 Refer to sections below for approved methods and other technical requirements: 

a) Almonds – AFL – Export to EU: See §18. 

b) Peanuts – AFL – U.S. Domestic & U.S. Imports: See §19. 
Agricultural Marketing Service

Science & Technology Program

Laboratory Approval and Testing Division

Laboratory Approval Service

Laboratory Approval Program – Mycotoxins

Doc. No: LAP-PR.07

Revision Date: 02/08/23 Page 8 of 29

c) Peanuts – AFL – Export to EU: See §20. 

d) Pistachios – AFL – U.S. Domestic & U.S. Imports: See §21. 

e) Pistachios – AFL – Export to EU: See §22. 

f) Pistachios – OTA – Export to EU: See §23. 

10 Demonstration of Method Performance by Validation and Verification Evaluation 

10.1 Demonstrate the sample test method(s) is competently and proficiently performed prior to 

using it for testing and reporting results by using a validation / verification process, where: 

a) Validation is the process of demonstrating that a method is suitable for its intended 

purpose, 

b) Verification is the process of demonstrating a validated method can be performed to the 

same level of performance determined during the validation, and 

c) It is recognized that there are multiple ways to demonstrate performance; therefore, 

where the Program does not specify a specific protocol, consult with the Program 

Manager to determine a suitable validation / verification plan using U.S. and 

internationally recognized protocols. 

10.2 Demonstrate that variations or modifications to a validated method (e.g., different matrix, 

different analyte, etc.) are fit-for-purpose. 

10.3 Prepare a Method Validation Data Package to record the procedure has been demonstrated 

as fit for use. 

a) Consolidate pertinent information into an integrated and auditable data package to 

support the objective’s conclusion that includes at least the following: cover statement 

containing the conclusion of the validation / verification process, test method

procedure, relevant statistics (i.e., linearity, accuracy, precision, measurement 

uncertainty, and limit of quantitation), and traceable data (raw and summarized). 

b) Send validation / verification package to the PM for review and approval. 

c) Ensure validation / verification data package is readily available at the laboratory for as 

long as the method is utilized, plus three years after the last reported results. 

d) Ensure official record of approval for method is maintained and readily available. 

10.4 Validate the sample preparation procedure for grinding samples to achieve an adequate 

particle size and homogenous grind for each analyte/matrix. 

a) Determine optimal parameters to achieve a homogeneous grind, including type of 

grinder, type of blade (e.g., smooth-edge, serrated, notch-edge), and minimum time to 

grind. 
Agricultural Marketing Service

Science & Technology Program

Laboratory Approval and Testing Division

Laboratory Approval Service

Laboratory Approval Program – Mycotoxins

Doc. No: LAP-PR.07

Revision Date: 02/08/23 Page 9 of 29

b) Select an unground sample with either a known incurred natural mycotoxins 

contamination or spike the nuts (ideally in one location). 

c) Grind sample using optimized parameters. 

d) Analyze at least 10 representative subsamples to determine if the resulting grind is 

homogeneous. 

e) Evaluate results from the subsamples. Acceptance criteria for a homogeneous grind is 

percent recovery as found in Table 1; and Coefficient of Variation (CV) ≤ 20%. 

f) Establish sieve test acceptance criteria for maintaining vertical cutter mill equipment by 

performing a particle size tests using subsamples of the grind passed through a No. 20 

sieve (see §14.3). 

g) Document, in a procedure, the final parameters validated including type of grinder, type 

of blade, minimum grind time, and sieve test acceptance criteria.

h) For significant changes made to the grinding procedure perform validation to 

demonstrate the changes result in a homogeneous grind and re-establish the sieve test 

acceptance criteria.

i) An exception or variation to AOAC 977.16 method may be considered by submitting a 

request and validation data that demonstrates the variation provides the same or better 

performance than the original procedure. 

10.5 Demonstrate the sample testing method(s) can be competently and proficiently performed 

prior to using it for testing and reporting results.

a) Demonstrate performance criteria of the reference method (e.g., AOAC) are met or 

exceeded (e.g., linearity, accuracy, precision, measurement uncertainty, limit of 

quantitation (lowest concentration that can be reliably detected and measured with 

accuracy and precision).

b) Demonstrate performance criteria outlined in EC 401/2006 of 23 February 2006 Annex II 

Part 4 are met, as relevant to scope of approval including export to EU (e.g., parts 4.2, 

4.3, 4.4, and 4.5) (see §5.8.a)). 

11 Analyst Competency and Proficiency Testing

11.1 Analyst Competency: 

a) Ensure new analyst responsible for performing test method(s) demonstrates 

competency and provide record of training and competence evaluation to the PM. 

b) Ensure every analyst responsible for performing the method(s) participates in the PT 

program at least once in a two-year span. 
Agricultural Marketing Service

Science & Technology Program

Laboratory Approval and Testing Division

Laboratory Approval Service

Laboratory Approval Program – Mycotoxins

Doc. No: LAP-PR.07

Revision Date: 02/08/23 Page 10 of 29

11.2 Laboratory Analytical Competency:

a) Participate in a PT program on a quarterly basis.

b) Use a PT program relevant to the approved method in terms of analyte, concentration 

range, and matrix on the scope of approval—administered by an external third-party 

program such as AOCS, FAPAS, or similar (with preference to ISO 17043 accredited

provider).

c) Participate in the FAPAS PT program when scope of approval includes export to EU.

d) Participate in PT Program and at the frequency specified by the Program Manager 

when a quarterly PT program is unavailable.

e) Review PT report and initiate the corrective action process when unsatisfactory results 

are observed. Unsatisfactory PT results and action to take are defined as follows:

|z| ≥ 3 Initiate immediate corrective action investigation on the part of the 

laboratory to establish root cause.

2 ≤ |z| ≤ 3 Evaluate in the context of other scores obtained in the same test and 

other PTs over time. Investigate to determine the root cause and take 

action as needed.

11.3 Submit to the PM:

a) Record of new analyst training and demonstration of competency.

b) Copy of the PT report with analyst name, and PT ID, within 30 days of receipt.

c) Record of corrective action response for unsatisfactory results, within 30 days of 

receiving the PT report.

*****

The following sections include requirements organized by scope; and are in 

addition to the requirements outlined above.
Agricultural Marketing Service

Science & Technology Program

Laboratory Approval and Testing Division

Laboratory Approval Service

Laboratory Approval Program – Mycotoxins

Doc. No: LAP-PR.07

Revision Date: 02/08/23 Page 11 of 29

TECHNICAL REQUIREMENTS (GENERAL) 

12 Performance Criteria

Table 1. Performance criteria for aflatoxins (B1, B2, G1, G2).

Criterion Concentration 

Range

Recommended 

Value

Maximum permitted 

Value

Blanks* All Negligible

Recovery* < 1.0 ug/kg 50 to 120%

Aflatoxins B1, B2, G1, G2 1-10 ug/kg 70 to 110%

> 10 ug/kg 80 to 110%

Reproducibility*

RSD All As derived from 

Horwitz Equation

2 x value derived from

Horwitz Equation

Repeatability*

RSDr

May be calculated as 0.66 times Reproducibility RSDR at the concentration 

of interest

Measurement Uncertainty**

Fit-for-use

Certainty of 95% at target concentration

Calculation Concentration Range Equation

Horwitz Equations† 1.2 xx 10−7 ≤ CC ≤ 0.138 RSDR = 2(1-0.5 log C)

CC < 1.2 xx 10−7 RSDR = 22%

Note: Values apply to B1 and sum of B1 + B2 + G1 + G2 

* EC 401/2006, Annex II, 4.3.1.1(a)

** EC 401/2006, Annex II, 4.2.2.2

† EC 401/2006, Annex II, 4.3.1.1(i)

Table 2. Performance criteria for ochratoxin A.

Level ug/kg* RSDr %* RSDR %* Recovery %* Measurement 

Uncertainty**

< 1 ≤ 40 ≤ 60 50 to 110 Certainty of 95% at 

≥ 1 ≤ 20 ≤ 30 70 to 110 target concentration

* EC 401/2006, Annex II, 4.3.1.1.b

** EC 401/2006, Annex II, 4.4.1

13 Quality Controls 

13.1 Use controls, daily with use, to demonstrate testing is performed correctly and factors that 

could negatively impact the results are mitigated.

13.2 Take immediate action prior to continuing testing and/or reporting results when any quality 

control does not perform as expected. 
Agricultural Marketing Service

Science & Technology Program

Laboratory Approval and Testing Division

Laboratory Approval Service

Laboratory Approval Program – Mycotoxins

Doc. No: LAP-PR.07

Revision Date: 02/08/23 Page 12 of 29

13.3 Include minimum quality controls:

a) Matrix spike, matrix blank and/or reagent blank, calibration standard, and 

b) For LC-FLD, or similar method, continued calibration verification (CCV). 

13.4 LAS’ interpretation of each quality control and its purpose is defined below. It is not a 

requirement for the laboratory to use the same term for each type of control as long as the 

correct control is used for the correct purpose. 

a) Matrix Blank: Substance that closely matches the samples being analyzed with regard 

to matrix components. Ideally, the matrix blank does not contain the analyte(s) of 

interest but is subjected to all sample processing operations including all reagents used 

to test the samples. Use to determine the absence of significant interference due to 

matrix, reagents, and equipment used in the analysis. 

b) Matrix Spike: Aliquot of a sample prepared by adding a known amount of analyte(s) to 

a specified amount of matrix and subjected to the entire procedure. Use to establish if 

the method is appropriate for the analysis of a specific analyte(s) in a particular matrix.

c) Incurred Sample: Sample that contains the analyte(s) of interest, which were not 

derived from laboratory fortification. 

d) Reagent Blank: Sample containing only the reagents taken through the entire 

procedure. Optional if a matrix blank is used. 

e) Calibration Standard: Solution containing known amount or concentration of analyte. 

Without further indication, calibration standard(s) is usually prepared by adding the 

analyte(s) of interest in neat solvent/solution. A “matrix-matched calibration 

standard(s)” is prepared by adding analyte(s) of interest in matrix blank. Use to 

establish a reference for identification and/or quantification for measuring/detection 

system.

f) Continuing Calibration Verification (CCV): Solution of known concentration, typically 

at or near the midpoint of the calibration curve. Use to evaluate instrument stability

throughout the sequence and repeatability. 

g) Independent Calibration Verification (ICV): Solution of known concentration that is 

from a different manufacturer, or same manufacturer but different lot, or a separate 

preparation from the solutions used to calibrate. Use to evaluate the accuracy of 

reference material and/or accuracy of preparation techniques. Optional; however, 

inclusion is a best practice.

14 Quality Measures

14.1 Evaluate quality controls to identify acceptability of data, trends, and potential problems. 

14.2 Take immediate action prior to continuing testing and/or reporting results when any quality 

measure not performing as expected.
Agricultural Marketing Service

Science & Technology Program

Laboratory Approval and Testing Division

Laboratory Approval Service

Laboratory Approval Program – Mycotoxins

Doc. No: LAP-PR.07

Revision Date: 02/08/23 Page 13 of 29

14.3 Vertical Cutter Mill (VCM) Sieve Test: Use to monitor VCM equipment (i.e., blade 

sharpness). Perform at a minimum frequency of weekly or with use.

a) Use a No. 20 sieve.

b) Place ~50 - 100g of ground sample in sieve that has been weighed.

c) Work the sample through the sieve with warm water (tap) being careful not to lose 

sample, as needed.

d) Dry the sieve and remaining material in an oven at ~100 – 105oC or air-dry until there 

is no more change in weight. Avoid cooking or burning the material.

e) Weigh and calculate % pass through.

f) Record result and evaluate against acceptance criteria defined in sample preparation 

procedure.

g) Monitor trending decrease of % pass through to facilitate maintaining acceptable VCM 

performance.

h) Submit sieve test data to the PM quarterly.

14.4 Calibration Curve: Use to quantify concentration of unknown samples.

a) Use the same type of curve used during method validation.

b) Ensure a minimum of 4 points is used for LC.

c) Ensure R2 value for each curve must be ≥0.995.

d) Ensure regression equation is not forced through zero.

e) Ensure calibration range brackets the range of result reported and includes the LOQ for 

the method. Results outside of the calibration range may be diluted and re-analyzed so 

the result falls within the range. It is not acceptable to extrapolate the concentration.

14.5 Continuing Calibration Verification (CCV): Use to demonstrate repeatability and stability 

of the instrument/method (e.g., calibration curve).

a) Collect CCV data points at a minimum, at the beginning of a batch, the end of the 

batch, and throughout the batch with a recommended frequency of every 10 (± 2) 

injections.

b) Calculate percent recovery of CCV data point (see §14.7).

c) Evaluate against the acceptance criteria where percent recovery must within 15% of 

expected concentration.

14.6 Coefficient of Variation (CV): Use to demonstrate the repeatability.

a) Calculate coefficient of variation (CV) for a group of data points using equation:

% CCCC = SSSSSSSSSSSSSSSS DDDDDDDD DDDD

AA AA ∗ 100
Agricultural Marketing Service

Science & Technology Program

Laboratory Approval and Testing Division

Laboratory Approval Service

Laboratory Approval Program – Mycotoxins

Doc. No: LAP-PR.07

Revision Date: 02/08/23 Page 14 of 29

b) Evaluate % CV during validation of sample preparation procedure (see §10.4); and 

during validation of sample testing method (see §10.5). 

14.7 Percent recovery: Use to compare expected concentration to actual measured concentration 

to evaluate method performance. 

a) Calculate percent recovery using equation: 

% RR RR RR = 

[ ffll ]

[ ee ] ∗ 100

b) Evaluate recovery of a matrix spike for aflatoxins against performance criteria defined 

in Table 1 (see §12). 

c) Evaluate recovery of a matrix spike for ochratoxin A against performance criteria 

defined in Table 2 (see §12).

14.8 Control Charting: Use to track performance over time and serve as an indicator if 

something in the analytical process is out of control and needs investigation or correction. 

a) Plot percent recovery results of each matrix spike analyzed. 

b) Plot for the method regardless of analyst or instrument used to evaluate overall 

performance of the method.

c) Evaluate whether results are acceptable using acceptance criteria, Table 1 (AFL) or 

Table 2 (OTA). All data points must be within the acceptable range.

d) Additional, optional, control charting methods can be used to evaluate other variables 

of the testing procedure. For example, plotting by analyst or by instrument can show 

individual performance when more than one is used. 

15 Data Analysis 

15.1 Significant Figures: Collect data to the appropriate significant figures for the methodology 

(see §18-23). Additionally, when two or more data points are calculated, the final value 

must not have more significant figures than the original data points. 

15.2 Rounding Rule: If 4 and under, round down; and if 5 and over, round up. For example:

a) a result of 9.4 ppb would be rounded to 9 ppb, whereas a result of 9.5 ppb would be 

rounded to 10 ppb for the final report; and

b) a result of 4.24 ppb would be rounded to 4.2 ppb, whereas a result of 4.25 ppb would be 

rounded to 4.3 ppb. 

15.3 Correction for recovery (export only): Correct the analytical result for recovery,

a) unless the percent recovery is between 90-110%, additionally
Agricultural Marketing Service

Science & Technology Program

Laboratory Approval and Testing Division

Laboratory Approval Service

Laboratory Approval Program – Mycotoxins

Doc. No: LAP-PR.07

Revision Date: 02/08/23 Page 15 of 29

b) the regulation states that if the result is less than 50% of the maximum level or 5 times 

the maximum level it might not be reported.

§ 4.4.1(a), Annex II, EC 401/2006, “Corrected for recovery, the level of recovery being 

indicated. The correction for recovery is not necessary in case the recovery rate is 

between 90-110 %.”

§ 4.4.1, Annex II, EC 401/2006, “if the result of the analysis is significantly (>50%) 

lower than the maximum level or much higher than the maximum level (i.e., more than 

5 times the maximum level), and on the condition that the appropriate quality 

procedures are applied and the analysis serves only the purpose of checking compliance 

with legal provisions, the analytical result might be reported without the correction for 

recovery and the reporting of the recovery rate and measurement uncertainty might be 

omitted in these cases.”

16 Critical Equipment & Reagents

16.1 Calibrate or verify equipment and instrumentation considered critical to the analytical 

method before putting into service and thereafter calibrate / verify regularly while in 

service.

a) Calibration refers to checking the measurements of a device and adjusting the device if 

corrections are needed.

b) Verification refers to checking that a device’s measurement remains within a 

determined acceptable range, adjustments should not be needed. 

16.2 When a device is found to be outside tolerance at verification, investigate and correct issue 

before using. 

16.3 Maintain records of calibration, intermediate checks, performance verification, 

maintenance, and significant repair.

16.4 Grinding Equipment: Keep in good working order. Monitor performance using VCM 

sieve test (See §14.3). Record blade sharpening. 

16.5 Balances & Scales:

a) Daily when in use: Verify with working weights that fall within the weight range used 

by the laboratory. 

b) Annual calibration: Performed by an accredited organization or internally with the 

same requirements being met.

16.6 Reference & Working Weights:

a) Reference weights: Verify every 5 years.

b) Working weights: Verify against reference weights annually.
Agricultural Marketing Service

Science & Technology Program

Laboratory Approval and Testing Division

Laboratory Approval Service

Laboratory Approval Program – Mycotoxins

Doc. No: LAP-PR.07

Revision Date: 02/08/23 Page 16 of 29

16.7 Analytical Instruments: Keep in good working order.

16.8 Volumetric Devices:

a) Mechanical pipets, micropipettors, mechanical burets, and bottle-top dispensers: 

Verify at least every 6 months for accuracy and precision using a gravimetric or 

colorimetric method.

b) Positive displacement syringes: Verify for accuracy upon receipt (a manufacturer’s 

certificate of accuracy may be accepted).

c) Plastic graduated cylinders: Verify for accuracy and precision upon receipt (prior to 

use) and every five years for accuracy using a gravimetric method. Additional 

verification should occur if damage is visible. 

d) Volumetric non-class A glassware: Verify for accuracy and precision using a 

gravimetric method upon receipt (prior to use). 

16.9 Critical reagents and consumables: Store according to the manufacturer’s instructions and 

do not use past the designated expiration date. 

17 Official Certificate of Analysis/Report

17.1 Meet the ISO 17025 standard for reporting results. 

17.2 Meet commodity specific requirements for reporting results. 

17.3 Meet customer’s requirements for reporting results. 

***** 

The following sections include requirements organized by scope; and are in 

addition to the requirements outlined above. 
Agricultural Marketing Service

Science & Technology Program

Laboratory Approval and Testing Division

Laboratory Approval Service

Laboratory Approval Program – Mycotoxins

Doc. No: LAP-PR.07

Revision Date: 02/08/23 Page 17 of 29

TECHNICAL REQUIREMENTS (SCOPE SPECIFIC)

18 Almond – AFL – Export to EU 

18.1 Applies to analysis for aflatoxins (B1, B2, G1, and G2). 

18.2 Sample Preparation Method: 

a) Protect sample from daylight (See §5.8.a), Annex II 1.1). 

b) Use a sample preparation procedure based on AOAC 977.16 to achieve a minimum size 

reduction for ground samples (See §10 and §24). 

c) Ensure sample is representative of the incoming sample prior to grinding. Mixing may 

occur at 1) the processing facility as part of the sampling process (See §5.6.b), Product 

Sample Selection and Analysis); or 2) at the laboratory (See §5.8.a), Annex I, D.2.4). 

When the incoming sample is split prior to grinding at the laboratory, ensure the subsamples are representative of the incoming sample.

18.3 Test Method: 

a) IAC with LC-FLD, Iodine: AOAC 991.31, A-F, H 

b) IAC with LC-FLD, Kobra Cell: AOAC 991.31, A-F, H / AOAC 999.07 

c) IAC with LC-FLD, PHRED: AOAC 991.31, A-F, H / AOAC 2005.08 

18.4 Data Analysis: 

a) Analyze to the nearest tenth ppb (i.e. 0.1 ppb) where ppb = 1 ng/g. 

b) Calculate and report shell/kernel ratio of whole nuts (See §5.8.a), Annex II 1.2), if 

applicable. 

18.5 Official Certificate of Analysis / Test Report: 

18.5.1 Include all the following information: 

a) Sample lot number.

b) Analytical method used (see §5.8.a)). 

c) Limit of quantification (see §5.8.a)). 

d) Percent recovery for B1 and total aflatoxin (see §5.8.a)). 

e) Analytical result (see §5.8.a)). 

f) A statement about correction for recovery (see §5.8.a)). 

g) Measurement uncertainty presented as the result ± uncertainty (i.e., 10.4 ± 2.1 

μg/kg), 
Agricultural Marketing Service

Science & Technology Program

Laboratory Approval and Testing Division

Laboratory Approval Service

Laboratory Approval Program – Mycotoxins

Doc. No: LAP-PR.07

Revision Date: 02/08/23 Page 18 of 29

§ 4.4.1(b), Annex II, EC 401/2006, “As x ± U whereby x is the analytical result and 

U is the expanded measurement uncertainty, using a coverage factor of 2 which 

gives a level of confidence of approximately 95%.” 

18.5.2 If a statement is made concerning compliance with the EC 401/2006, use the statement:

“Sample analysis was conducted in compliance with Annex II of Regulation (EC) No.

401/2006 for aflatoxin in almonds.” 

18.5.3 If reporting results for in shell nuts use the statement (as applicable):

“In shell almonds are reported on a kernel basis for a [insert X/Y] shell/kernel ratio.”

18.5.4 If requested by the handler to include an interpretation statement on the analytical 

report of the maximum limits set by Regulation EC 1881/2006, use the following types 

of statements. 

Note: Interpretation of results by the laboratory does not generally occur. The 

commodity’s destination is often decided based on the test results. It is the 

responsibility of the handler (processor) not the laboratory, to determine which limits 

apply to their product. 

a) “result exceeds the limit of 12.0 μg/kg aflatoxin B1 and/or 15.0 μg/kg sum of 

aflatoxin B1, B2, G1 and G2 for almonds to be subjected to sorting, or other physical 

treatment, before human consumption or use as an ingredient in foodstuffs (§2.1.2 

of Annex II Regulation EC 1881/2006).” 

b) “result exceeds the limit of 8.0 μg/kg aflatoxin B1 and/or 10.0 μg/kg sum of 

aflatoxin B1, B2, G1 and G2 for almonds intended for direct human consumption or 

use as an ingredient in foodstuffs (§2.1.6 of Annex II Regulation EC 1881/2006).” 
Agricultural Marketing Service

Science & Technology Program

Laboratory Approval and Testing Division

Laboratory Approval Service

Laboratory Approval Program – Mycotoxins

Doc. No: LAP-PR.07

Revision Date: 02/08/23 Page 19 of 29

19 Peanut – AFL – U.S. Domestic & U.S. Imports 

19.1 Applies to analysis for aflatoxins (B1, B2, G1, and G2). 

19.2 Sample Preparation Method: 

a) Visually inspect to verify incoming sample bag (unground) is marked with weight on 

the positive lot identification (PLI) tag (See §5.4.e), Sample Weight) and it is not less 

than 48lbs. 

b) Use sample preparation procedure based on AOAC 977.16 to achieve a minimum size 

reduction for ground samples (See §10 and §24). 

c) Ensure sample is representative of the incoming sample prior to grinding. 

Note: Samples are mixed prior to grinding at the processing facility as part of the 

USDA Federal/State Inspection Service sampling process (See §5.4.e), Sampling). 

19.3 Test Method: 

a) IAC with direct fluorometry: AOAC 991.31, A-G 

b) TLC: AOAC 998.03 

c) IAC with LC-FLD, Iodine: AOAC 991.31, A-F, H 

d) IAC with LC-FLD, Kobra Cell: AOAC 991.31, A-F, H / AOAC 999.07 

e) IAC with LC-FLD, PHRED: AOAC 991.31, A-F, H / AOAC 2005.08 

19.4 Data Analysis: 

19.4.1 Analyze to the nearest: 

a) IAC with direct fluorometry: whole integer 

b) TLC: whole integer 

c) IAC with LC-FLD: tenth ppb (i.e. 0.1 ppb) where ppb = 1 ng/g 

19.5 Official Certificate of Analysis / Test Report: 

19.5.1 Include the statement: 

“The designation of aflatoxin negative is defined as the average analytical result of 15 

parts per billion (ppb) or less aflatoxin and applies to product distributed within the 

United States under 7 CFR Part 996. Results are reported as whole integers; therefore, 

further calculations must be rounded to the nearest whole integer for proper 

interpretation.” 

19.5.2 Include the statement: 

“USDA or USDA-approved laboratory to test for total aflatoxin content in samples for 

domestic and imported peanuts marketed in the United States.” 
Agricultural Marketing Service

Science & Technology Program

Laboratory Approval and Testing Division

Laboratory Approval Service

Laboratory Approval Program – Mycotoxins

Doc. No: LAP-PR.07

Revision Date: 02/08/23 Page 20 of 29

19.5.3 Include one of the following statements for the methodology as approved, to include 

AOAC method number or internal procedure number, where applicable:

a) Immunoaffinity column with direct fluorometry method of analysis (AOAC 

991.31). 

b) Water slurry method with thin-layer chromatography (TLC) analysis designated as 

the alternative Best Foods (BF) method of analysis (AOAC 998.03). 

c) Immunoaffinity column cleanup with high performance liquid chromatography 

(LC) method. 

19.6 Reporting to USDA: 

a) Submit records of sample analysis to AMS Specialty Crops Inspection Division 

(sciinspectionoperations@usda.gov) in the format requested. 

b) Submit records of sample analysis to AMS Specialty Crops Market Development

Division (8Eimports@usda.gov and complianceinfo@usda.gov) in the format 

requested. 
Agricultural Marketing Service

Science & Technology Program

Laboratory Approval and Testing Division

Laboratory Approval Service

Laboratory Approval Program – Mycotoxins

Doc. No: LAP-PR.07

Revision Date: 02/08/23 Page 21 of 29

20 Peanut – AFL – Export to EU 

20.1 Applies to analysis for aflatoxins (B1, B2, G1, and G2). 

20.2 Sample Preparation Method: 

a) Visually inspect to verify incoming unground sample has the weight recorded on the 

positive lot identification (PLI) tag and it is not less than 48lbs (See §5.4.e), Sample 

Weight). 

b) Protect sample from daylight (See §5.8.a), Annex II 1.1).

c) Use sample preparation procedure based on AOAC 977.16 to achieve a minimum size 

reduction for ground samples (See §10 and §24). 

d) Ensure the sub-samples are representative of the incoming sample when the incoming 

sample is split prior to grinding at the laboratory. 

Note: Samples are mixed prior to grinding at the processing facility as part of the 

USDA Federal/State Inspection Service sampling process (See §5.4.e), Sampling). 

20.3 Test Method: 

a) IAC with LC-FLD, Iodine: AOAC 991.31, A-F, H

b) IAC with LC-FLD, Kobra Cell: AOAC 991.31, A-F, H / AOAC 999.07 

c) IAC with LC-FLD, PHRED: AOAC 991.31, A-F, H / AOAC 2005.08 

20.4 Data Analysis: 

a) Analyze to the nearest tenth ppb (i.e. 0.1 ppb) where ppb = 1 ng/g. 

b) Calculate and report shell/kernel ratio of whole nuts (See §5.8.a), Annex II 1.2), if 

applicable. 

20.5 Official Certificate of Analysis / Test Report: 

20.5.1 Include all the following information: 

a) Sample lot number (positive lot identification, PLI).

b) Analytical method used (see §5.8.a)). 

c) Limit of quantification (see §5.8.a)). 

d) Percent recovery for B1 and total aflatoxin (see §5.8.a)). 

e) Analytical result (see §5.8.a)). 

f) A statement about correction for recovery (see §5.8.a)). 

g) Measurement uncertainty presented as the result ± uncertainty (i.e., 10.4 ± 2.1 

μg/kg), 
Agricultural Marketing Service

Science & Technology Program

Laboratory Approval and Testing Division

Laboratory Approval Service

Laboratory Approval Program – Mycotoxins

Doc. No: LAP-PR.07

Revision Date: 02/08/23 Page 22 of 29

§ 4.4.1(b) Annex II, EC 401/2006, “As x ± U whereby x is the analytical result and 

U is the expanded measurement uncertainty, using a coverage factor of 2 which 

gives a level of confidence of approximately 95%.” 

h) State whether the sheller is a signatory of the APC MOU.

20.5.2 If a statement is made concerning compliance with the EC 401/2006, use the statement:

“Sample analysis was conducted in compliance with Annex II of Regulation EC No. 

401/2006 for aflatoxin in peanuts.” 

20.5.3 If reporting results for in shell nuts use the statement (as applicable):

“In shell peanut are reported on a kernel basis for a [insert X/Y] shell/kernel ratio.”

20.5.4 If requested by the handler to include an interpretation statement on the analytical 

report of the maximum limits set by Regulation EC 1881/2006, use the following types 

of statements. 

Note: Interpretation of results by the laboratory does not generally occur. The 

commodity’s destination is often decided based on the test results. It is the 

responsibility of the handler (processor) not the laboratory, to determine which limits 

apply to their product. 

a) “result exceeds the limit of 8.0 μg/kg aflatoxin B1 and/or 15.0 μg/kg sum of 

aflatoxin B1, B2, G1 and G2 for peanuts to be subjected to sorting, or other physical 

treatment, before human consumption or use as an ingredient in foodstuffs with the 

exception of peanuts for crushing for refined vegetable oil production (§2.1.1 of 

Annex II Regulation EC 1881/2006).” 

b) “result exceeds the limit of 2.0 μg/kg aflatoxin B1 and/or 4.0 μg/kg sum of aflatoxin 

B1, B2, G1 and G2 for peanuts intended for direct human consumption or use as an 

ingredient in foodstuffs with the exception of crude vegetable oils destined for 

refining and refined vegetable oils (§2.1.5 of Annex II Regulation EC 1881/2006).” 
Agricultural Marketing Service

Science & Technology Program

Laboratory Approval and Testing Division

Laboratory Approval Service

Laboratory Approval Program – Mycotoxins

Doc. No: LAP-PR.07

Revision Date: 02/08/23 Page 23 of 29

21 Pistachio – AFL – U.S. Domestic & U.S. Imports

21.1 Applies to analysis for aflatoxins (B1, B2, G1, and G2). 

21.2 Sample Preparation Method: 

a) Use sample preparation procedure based on AOAC 977.16 to achieve a minimum size 

reduction for ground samples (See §10 and §24). 

b) Ensure sample is representative of the incoming sample prior to grinding. Mixing may 

occur at 1) the processing facility as part of the sampling process (See §5.4.g) and 

5.4.h)); or 2) at the laboratory. 

21.3 Test Method: 

a) IAC with direct fluorometry: AOAC 991.31, A-G 

b) IAC with LC-FLD, Iodine: AOAC 991.31, A-F, H 

c) IAC with LC-FLD, Kobra Cell: AOAC 991.31, A-F, H / AOAC 999.07 

d) IAC with LC-FLD, PHRED: AOAC 991.31, A-F, H / AOAC 2005.08 

21.4 Data Analysis: 

a) Analyze to the nearest tenth ppb (i.e. 0.1 ppb) where ppb = 1 ng/g. 

21.5 Official Certificate of Analysis / Test Report: 

21.5.1 Include the statement: 

“USDA or USDA-approved laboratory to test for total aflatoxin content in samples for 

domestic and imported pistachios marketed in the United States.” 

21.5.2 Include one of the following statements for the methodology used, to include AOAC 

method number or internal procedure number, where applicable: 

a) Immunoaffinity column with direct fluorometry method of analysis (AOAC 

991.31). 

b) Immunoaffinity column cleanup with high performance liquid chromatography 

(LC) method.

21.5.3 If requested by the handler to include an interpretation statement on the analytical 

report of the maximum, use the following types of statements. 

Note: Interpretation of results by the laboratory does not generally occur. The 

commodity’s destination is often decided based on the test results. It is the 

responsibility of the handler (processor) not the laboratory, to determine which limits 

apply to their product. 

a) “result exceeds the limit of 15 ppb total aflatoxin for domestic human consumption 

(7 CFR 983.150).” 
Agricultural Marketing Service

Science & Technology Program

Laboratory Approval and Testing Division

Laboratory Approval Service

Laboratory Approval Program – Mycotoxins

Doc. No: LAP-PR.07

Revision Date: 02/08/23 Page 24 of 29

21.6 Reporting to USDA: 

a) Submit records of sample analysis to AMS Specialty Crops Inspection Division 

(sciinspectionoperations@usda.gov) in the format requested. 

b) Submit records of sample analysis to AMS Specialty Crops Market Development 

Division (8Eimports@usda.gov and complianceinfo@usda.gov) in the format 

requested. 
Agricultural Marketing Service

Science & Technology Program

Laboratory Approval and Testing Division

Laboratory Approval Service

Laboratory Approval Program – Mycotoxins

Doc. No: LAP-PR.07

Revision Date: 02/08/23 Page 25 of 29

22 Pistachio – AFL – Export to EU

22.1 Applies to analysis for aflatoxins (B1, B2, G1, and G2). 

22.2 Sample Preparation Method: 

a) Protect sample from daylight (See §5.8.a) Annex II 1.1).

b) Use sample preparation procedure based on AOAC 977.16 to achieve a minimum size 

reduction for ground samples (See §10 and §24). 

c) Ensure sample is representative of the incoming sample prior to grinding. Mixing may 

occur at 1) the processing facility as part of the sampling process (See §5.6.c), Creating 

and Sampling the Lot); or 2) at the laboratory (See §5.8.a), Annex I, D.2.4). When the 

incoming sample is split prior to grinding at the laboratory, ensure the sub-samples are 

representative of the incoming sample.

22.3 Test Method: 

a) IAC with LC-FLD, Iodine: AOAC 991.31, A-F, H

b) IAC with LC-FLD, Kobra Cell: AOAC 991.31, A-F, H / AOAC 999.07 

c) IAC with LC-FLD, PHRED: AOAC 991.31, A-F, H / AOAC 2005.08 

22.4 Data Analysis: 

a) Analyze to the nearest tenth ppb (i.e. 0.1 ppb) where ppb = 1 ng/g. 

b) Calculate and report shell/kernel ratio of whole nuts (See §5.8.a), Annex II 1.2), if 

applicable. 

c) In shell pistachio ratio is accepted as 50/50 (see §5.5.d)). 

22.5 Official Certificate of Analysis / Test Report: 

22.5.1 Include all the following information: 

a) Sample lot number.

b) Analytical method used (see §5.8.a)). 

c) Limit of quantification (see §5.8.a)). 

d) Percent recovery for B1 and total aflatoxin (see §5.8.a)). 

e) Analytical result (see §5.8.a)). 

f) A statement about correction for recovery (see §5.8.a)). 

g) Measurement uncertainty presented as the result ± uncertainty (i.e., 10.4 ± 2.1 

μg/kg). 

§ 4.4.1(b), Annex II, EC 401/2006, “As x ± U whereby x is the analytical result and 

U is the expanded measurement uncertainty, using a coverage factor of 2 which 

gives a level of confidence of approximately 95%.” 
Agricultural Marketing Service

Science & Technology Program

Laboratory Approval and Testing Division

Laboratory Approval Service

Laboratory Approval Program – Mycotoxins

Doc. No: LAP-PR.07

Revision Date: 02/08/23 Page 26 of 29

22.5.2 If a statement is made concerning compliance with the EC 401/2006, use the statement:

“Sample analysis was conducted in compliance with Annex II of Regulation EC No. 

401/2006 for aflatoxin in pistachio.” 

22.5.3 If reporting results for in shell nuts use the statement (as applicable):

“In shell pistachio results are reported on a kernel basis for a 50/50 shell/kernel ratio.” 

22.5.4 If requested by the handler to include an interpretation statement on the analytical 

report of the maximum limits set by Regulation EC 1881/2006, use the following types 

of statements. 

Note: Interpretation of results by the laboratory does not generally occur. The 

commodity’s destination is often decided based on the test results. It is the 

responsibility of the handler (processor) not the laboratory, to determine which limits 

apply to their product. 

a) “result exceeds the limit of 12.0 μg/kg aflatoxin B1 and/or aflatoxin 15.0 μg/kg sum 

of B1, B2, G1 and G2 for pistachios to be subjected to sorting, or other physical 

treatment, before human consumption or use as an ingredient in foodstuffs (§2.1.2 

of Annex II Regulation EC 1881/2006).” 

b) “result exceeds the limit of 8.0 μg/kg aflatoxin B1 and/or 10.0 μg/kg sum of 

aflatoxin B1, B2, G1 and G2 for pistachios intended for direct human consumption or 

use as an ingredient in foodstuffs (§2.1.6 of Annex II Regulation EC 1881/2006).” 
Agricultural Marketing Service

Science & Technology Program

Laboratory Approval and Testing Division

Laboratory Approval Service

Laboratory Approval Program – Mycotoxins

Doc. No: LAP-PR.07

Revision Date: 02/08/23 Page 27 of 29

23 Pistachio – OTA – Export to EU 

23.1 Applies to analysis of ochratoxin A. 

23.2 Sample Preparation Method:

a) Protect sample from daylight (See §5.8.a), Annex II 1.1).

b) Use sample preparation procedure based on AOAC 977.16 to achieve a minimum size 

reduction for ground samples (See §10 and §24). 

c) Ensure sample is representative of the incoming sample prior to grinding. Mixing may 

occur at 1) the processing facility as part of the sampling process (See §5.6.c), Creating 

and Sampling the Lot); or 2) at the laboratory (See §5.8.a), Annex I, D.2.4). When the 

incoming sample is split prior to grinding at the laboratory, ensure the sub-samples are 

representative of the incoming sample.

23.3 Test Method: 

a) Use method fit for purpose and that meets the validation / verification requirements 

outlined in §10. 

23.4 Data Analysis:

a) Analyze to the nearest tenth ppb (i.e. 0.1 ppb) where ppb = 1 ng/g. 

b) Calculate and report shell/kernel ratio of whole nuts (See §5.8.a), Annex II 1.2), if 

applicable. 

c) In shell pistachio ratio is accepted as 50/50 (see §5.5.d)). 

23.5 Official Certificate of Analysis / Test Report:

23.5.1 Include all the following information: 

a) Sample lot number.

b) Analytical method used (see §5.8.a)). 

c) Limit of quantification (see §5.8.a)). 

d) Percent recovery for OTA (see §5.8.a)). 

e) Analytical result (see §5.8.a)). 

f) A statement about correction for recovery (see §5.8.a)). 

g) Measurement uncertainty presented as the result ± uncertainty (i.e., 10.4 ± 2.1 

μg/kg). 

§ 4.4.1(b), Annex II, EC 401/2006, “As x ± U whereby x is the analytical result and 

U is the expanded measurement uncertainty, using a coverage factor of 2 which 

gives a level of confidence of approximately 95%.” 
Agricultural Marketing Service

Science & Technology Program

Laboratory Approval and Testing Division

Laboratory Approval Service

Laboratory Approval Program – Mycotoxins

Doc. No: LAP-PR.07

Revision Date: 02/08/23 Page 28 of 29

23.5.2 If a statement is made concerning compliance with the EC 401/2006, use the statement:

“Sample analysis was conducted in compliance with Annex II of Regulation (EC) No.

401/2006 for ochratoxin in pistachio.” 

23.5.3 If reporting results for in shell nuts use the statement (as applicable):

“In shell pistachio results are reported on a kernel basis for a 50/50 shell/kernel ratio.” 

23.5.4 If requested by the handler to include an interpretation statement on the analytical 

report of the maximum limits set by Regulation EC 1881/2006, use the following types 

of statements. 

Note: Interpretation of results by the laboratory does not generally occur. The 

commodity’s destination is often decided based on the test results. It is the 

responsibility of the handler (processor) not the laboratory, to determine which limits 

apply to their product. 

a) “result exceeds the limit of 10 μg/kg ochratoxin for pistachios to be subjected to 

sorting, or other physical treatment, before human consumption or use as an 

ingredient in foodstuffs (Annex II Regulation EC 1881/2006).” 

b) “result exceeds the limit of 5 μg/kg ochratoxin for pistachios intended for direct 

human consumption or use as an ingredient in foodstuffs (Annex II Regulation EC 

1881/2006).” 
Agricultural Marketing Service

Science & Technology Program

Laboratory Approval and Testing Division

Laboratory Approval Service

Laboratory Approval Program – Mycotoxins

Doc. No: LAP-PR.07

Revision Date: 02/08/23 Page 29 of 29

24 Explanations for Technical Requirements

24.1 For sample grinding: 

a) Particle size is a performance control measurement of homogeneity (See §5.7.f)). 

b) Adequate grind is described in AOAC 977.16 and summarized as: “Aim at maximum 

practical size reduction and thoroughness of mixing to achieve effective distribution of 

contaminated portions...To achieve this degree of size reduction, nut must be ground to 

pass No. 20 sieve. 

25 Revision History

New Rev. Description of Change Prepared by

08/12/22 Original. This document is the product of rebranding LAP-Aflatoxins to 

incorporate additional mycotoxins into the Program’s scope. 

Grace Vaillant

Branch Chief

Heath McClure

Program Manager

02/08/23 Added §20.5.1.h) “State whether the sheller is a signatory of the APC MOU”

because the requirement was added to the APC MOU for 2023.

Grace Vaillant

Branch Chief

26 Review / Approvals 

Heath McClure Grace Vaillant

Program Manager - Aflatoxin Branch Chief

Reviewer Approver