No safe blood lead concentration in children has been identified.^1,2 Lead can affect nearly every system in the body and is especially harmful to the developing central nervous systems of children.³. Chronic lead exposure may occur with no obvious symptoms, but it has been associated with developmental delay, sluggishness and fatigue, weight loss, irritability and difficulties learning.³ In 2012, the Centers for Disease Control and Prevention’s Advisory Committee on Childhood Lead Poisoning Prevention (ACCLPP) recommended using a population-based reference value, calculated as the 97.5^th percentile of blood lead in children one to five years old in the U.S., instead of a blood lead “level of concern” to identify children and environments associated with lead hazards.² Based upon the 2007–2010 National Health and Nutrition Examination Survey (NHANES) blood lead results, the reference value was 5 μg/dL. The ACCLPP recommended that CDC update the reference value every four years using the most recent NHANES blood lead data for children ages one to five years old.² CDC concurred or concurred in principle with the ACCLPP recommendations.⁴

Using available NHANES 2011–2014 data, we calculated the 97.5^th percentile at 3.48 μg/dL (95% confidence interval, 2.65, 4.29 μg/dL), approximately 30% lower than the current reference value. Our objective is to describe the laboratory implications of a decreasing trend in blood lead concentrations (referred to as blood lead levels, BLLs) in U.S. children and the clinical interpretation challenges that result from the variability of measurement of low BLLs. Because the CDC has not made a final decision about changing the current reference value of 5 μg/dL we refer to a calculated 97.5^th percentile rather than a reference value.

As BLLs in U.S. children have declined over time (see Figure), acceptability criteria for laboratory performance of blood lead analysis in proficiency testing (PT) programs have become more rigorous, requiring laboratories to change processes and technologies. Prior to 1992, PT performance was judged satisfactory if the laboratory reported results for test samples that were within ± 6 μg/dL or ± 15% of the assigned (target) concentration for individual PT samples.⁵ The Clinical Laboratory Improvement Amendments (CLIA) of 1988 tightened the acceptability criteria for blood lead measurements to ± 4 μg/dL or ± 10%, whichever is greater.⁶ In 2010, the ACCLPP recommended that the criteria be reduced to ± 2 μg/dL or ± 10%, whichever is greater,² noting that the majority of laboratories measuring blood lead were already achieving measurement errors of ± 2 μg/dL at these concentrations.⁷ However, the acceptability criteria in the CLIA regulations have not been updated.

Measuring ever-lower BLLs required laboratories to shift to newer technologies with lower method limits of detection (LODs) and improved accuracy and precision. Methodology in the 1970’s was based on flame absorption spectroscopy, later followed by methods based on electrothermal atomic absorption spectrometry, and anodic stripping voltammetry. In the 1990s inductively coupled plasma mass spectrometry (ICP-MS) was introduced and newer generations of ICP-MS have even higher sensitivity and lower background levels. These changes in analytical methodology have made it possible for laboratories to achieve lower LODs and make accurate, precise blood lead measurements significantly below 5 μg/dL. Nonetheless, achieving accurate and precise measurements at blood lead concentrations < 5 μg/dL is an analytical challenge using CLIA-exempt instruments (e.g., LeadCare^® II analyzer with an LOD of 3.3 μg/dL). In addition to using highly technical measurement methods, such as ICP-MS, special precautions are needed to avoid external lead contamination of the blood collection devices and instrument reagents.

The CDC’s Environmental Health Laboratory in the Division of Laboratory Sciences (EHL-DLS) plays an important public health role in blood lead measurement that includes NHANES sample analysis that serve as reference values for U.S. children and adults. Lot screening has been an important activity used by CDC’s laboratory to exclude entire lots of items used in blood collection (e.g., butterfly needles, collection tubes, alcohol and iodine wipes) or analytical laboratory materials (e.g., pipette tips, reagents) with unacceptable lead contamination that could result in falsely elevated BLLs. In addition, the CDC’s ICP-MS analytical method timing, calibrator placement, calibration regression type, sample introduction system, and reagent composition were optimized for accurate and precise determination of lead in blood at low concentrations. By employing all these measures, the EHL-DLS achieved a blood lead LOD of 0.07 μg/dL for the 2013–2014 NHANES survey period.

As BLLs in U.S. children continue to decrease, more laboratories will need to be able to accurately measure concentrations below their current LODs. To accomplish this, laboratories will need to consider various modifications, including selecting the optimal analytical method as well as testing for lead contamination of laboratory reagents and supplies used in the laboratory (e.g., aliquotting devices, cryovials). Lead contamination also may occur during blood sample collection because of external skin contamination and small amounts of lead in the blood collections materials (e.g., needles, vials, anticoagulants in tubes). Precautions to avoid skin contamination during blood collection are well known to clinicians and clinics that conduct lead testing. However, manufacturers of blood collection devices may need to consider screening devices for even lower levels of lead contamination that interfere with laboratory measurements and take actions to prevent lead contamination during production.

Although each device, reagent, or item that has contact with a child’s blood may have only a small amount of lead, these sources are additive to a blood sample throughout the pre-analytical and analytical process. Such contamination may have little impact to measurements of blood lead at values of 10 μg/dL or more; however, the sum of contamination sources may contribute significantly to blood lead measurements near or below the current reference value. It is a principle for blood lead or any analytical measurement that as the measurement approaches the LOD, the variability around the measurement increases significantly.⁸ Therefore, as BLLs continue to decline, laboratories may need to lower their analytical LODs using analytical process improvements, technology changes, or both. We describe several approaches used by the EHL-DLS to improve analytical accuracy, precision, and to reduce lead contamination during blood collection and specimen analysis.

BLLs in U.S. children one to five years old have declined (see Figure) to a point that challenges the detection limit of many laboratories. Children with BLLs at or above the 90^th or 97.5^th percentile for NHANES 2011–2014 have characteristics similar to what has been reported in past studies of risk factors: less than three years old; male; non-Hispanic black; and living in low income households. This suggests that lead-based paint hazards continue to be a source of childhood lead exposure, but we did not have geographic details to determine residence location or housing age.¹⁵ We could not evaluate sources of exposure and contributions from other sources, including contaminated soil, dust, drinking water, and occasional sources such as cosmetics, remedies, hobbies, and occupational take-home. Although the NHANES 2011–2014 sample of one to five year old children was large, the 97.5^th percentile and higher was comprised of only 46 children. This contributes to the wide variability around 3.48 μg/dL, with a 95% confidence interval of 2.65 to 4.29 μg/dL. Despite this limitation, NHANES is the best and possibly only data source for U.S. population-based estimates.

BLLs of 5 μg/dL also present a clinical interpretation challenge. Although reported accuracy for most laboratories is ± 2 μg/dL (Parsons et al, 2001), the current CLIA acceptability criteria for accuracy in blood lead measurements of ±4 μg/dL (at values less than 40 μg/dL) which means that the true value of a blood lead reported as 4 μg/dL can be between 0 and 8 μg/dL. Therefore, when the child is retested, any result between 0 and 8 μg/dL includes the possibility that the true BLL is unchanged. It would be helpful for the clinician to explain to a parent or guardian the concept of variability in the measurement if, for example, a child’s BLL goes from 4 to 8 μg/dL in the absence of a new or increased exposure.

Since almost 23% of LAMP-participating laboratories reported LODs between 3 and 5 μg/dL, it is likely that many laboratories will be unable to quantify blood lead at or near the 97.5^th percentile value (Table 5b). Forty percent of LAMP- participating laboratories were unable to quantify BLLs at around 1.5 μg/dL, implying that surveillance data collected from clinical laboratories for the general population could be at or below the LODs of many laboratories. So that LAMP participants can improve precision and accuracy of blood lead measurements below 5 μg/dL and to assist laboratories in testing new technology, CDC will include more challenge samples with target blood lead values between 1 and 5 μg/dL in future performance challenges.

Manufacturers of items such as blood collection materials and containers, cryovials, and reagents need increased awareness and to consider actions that avoid potential lead contamination during production of these items. Screening of blood collection devices is not feasible for most laboratories that provide blood lead measurements because they do not typically provide the blood collection materials. The EHL-DLS is finding it increasingly difficult to purchase manufactured lots that pass screening, and we anticipate that the percentage of device and reagent failures is likely to increase unless changes are made in manufacturing processes.

A tightening of the blood lead acceptability criteria in proficiency testing to ±2 μg/dL (≤20 μg/dL) or ± 10% from ±4 μg/dL (≤40 μg/dL) or ± 10% will encourage laboratories to be aware of, and to proactively deal with contamination and measurement issues that often plague the analysis of blood lead at low levels. If laboratories are able to reduce contamination associated with their measurements, the limits of detection should improve. Currently, 33% of U.S. LAMP participating laboratories report a blood lead limit of detection ≥ 2 μg/dL (the 2013–2014 90^th percentile for blood lead is 1.8 μg/dL). CDC will assist in reinforcing the need for blood lead laboratories to improvements contamination and measurement issues through the LAMP program. In 2017, LAMP reports will include telling participating laboratories how they would perform if a ±2 μg/dL or ±10% acceptance criteria were used.

The 97.5^th percentile BLL based upon NHANES 2011–2014 results in children ages one to five years is 3.48 μg/dL, 30% lower than the current reference value of 5 μg/dL. Although the number of children in the sample that comprised the 97.5^th percentile was small, they demonstrated previously identified risk factors for elevated BLLs: younger than three years old; male; non-Hispanic black; and living in low-income households. The continued decrease of BLLs in children presents challenges for clinicians, laboratories, and manufacturers of consumables and analytical instruments. To achieve precise and accurate blood lead measurements with lower limits of detection, laboratories need to evaluate potential sources of external lead contamination, optimize their analytical methods for low concentration measurements, and participate in external proficiency testing programs, considering how they would perform if tighter acceptability criteria were used.

Manufacturers of devices used in blood lead sample collection could identify potential sources of lead contamination and take actions to reduce these sources. Clinicians should understand the factors affecting accurate measurements at very low blood lead concentrations to better interpret BLLs and assess whether small changes are real or indicate measurement variability.