The 24-Hour Urine Collection component was added to NHANES in 2014 to: 1)
estimate dietary intakes of sodium and potassium; 2) estimate the excretion of
microalbumin (albumin and creatinine); and 3) assess measures of kidney markers
(phosphorus, magnesium, calcium, oxalate, and urea nitrogen), caffeine, volatile
organic compounds, and iodine in the U.S. population.
In 2014, a random half sample of non-pregnant participants aged 20-69 years
and examined in the mobile examination center (MEC) were asked to collect their
urines for a 24-hour period. For the first half of the year, a random half of
those who completed the initial 24-hour urine collection were recruited to
collect a second 24-hour urine specimen 3 to 10 days later. For the second half
of the year, all those who completed the first 24-hour urine collection were
recruited to collect a second 24-hour urine specimen. Data processing
information from the initial and second 24-hour urine collections are released
in two separate datasets (UR1_H_R and UR2_H_R). Each of these two files contain
information on: total urine volume, length of collection, completeness of the
urine collection, number of complete urine specimens, collection day of the
week, and responses to questions on the participants’ experience collecting the
urine to assess completeness of the specimen. Please see the documentation for
these two files for more details.
Separate datasets were produced to include laboratory results of analytes
from the 24-hour urine collections. The present file contains analyte data for
urine kidney markers (albumin, creatinine, calcium, magnesium, oxalate,
phosphorus, and urea nitrogen). See Appendix I for the list of data files for
the 24-hour urine collection.
Urinary kidney markers are a diverse set of analytes often used to measure
many complex and serious medical conditions including: obesity, renal disease,
cardiac disease, hypertension, diabetes and hepatic disease. Each analyte
provides unique information that may provide greater insight into many of the
challenging public health issues that are prevalent in the U.S. Additionally,
some of these kidney markers also aid in the assessment of nutritional status.
When used in conjunction with measurements of dietary sodium over a 24-hour
period, kidney marker measurements may aid in providing enhanced understanding
of how to efficiently address many of these growing public health concerns.
To reduce the risk of inadvertent disclosure, all data from this 1-year
24-hour urine collection can only be accessed through the NCHS Research Data
Center (RDC). Instructions for requesting use of these data are available at the
RDC website (https://www.cdc.gov/rdc/).
The eligible sample was a random one-half sample of all examined participants
aged 20-69 years with a few participants eliminated based on exclusion criteria
(e.g. pregnant). Among participants with a complete 24-hour urine specimen, a
portion of them were eligible for collection of a second 24-hour urine specimen.
For the first half of the year, a random one-half were asked to collect a second
specimen. For the second half of the year, all of the eligible participants were
asked to collect a second 24-hour urine specimen.
Description of Laboratory
A fluorescent immunoassay for the measurement of human urinary albumin is
described by Chavers et al. The methodology involves solid-phase,
non-competitive, double-antibody reaction. Urine specimen albumin antigen reacts
with albumin antibody that is covalently attached to polyacrylamide beads. This
resulting solid-phase antibody complex is then reacted with fluorescein-labeled
antibody. Unattached fluorescent antibody and other proteins are removed by
centrifugation. The fluorescence of the stable solid-phase double-antibody
complex is measured with a fluorometer and is directly proportional to the
amount of urine albumin present. The standard line calibration material is
human serum albumin with a range of 0.5 to 20 µg/mL.
In this enzymatic method, creatinine is converted to creatine under the
activity of creatininase. Creatine is then acted upon by creatinase to form
sarcosine and urea. Sarcosine oxidase converts sarcosine to glycine and hydrogen
peroxide, and the hydrogen peroxide reacts with chromophore in the presence of
peroxidase to produce a color product that is measured at 546 nm (secondary
wavelength = 700 nm). This is an endpoint reaction that agrees well with
recognized HPLC methods, and it has the advantage over Jaffe picric acid-based
methods that are susceptible to interferences from non-creatinine
This method utilizes ammonium molybdate as the color-forming reagent.
Measurement of the final product occurs at 340 nm (secondary wavelength 700 nm).
Inorganic phosphate forms an ammonium phosphomolybdate complex having the
formula (NH4)3[PO4 (MoO3)12] with ammonium molybdate in the presence of sulfuric
acid. The concentration of phosphomolybdate formed is directly proportional to
the inorganic phosphate concentration.
In this method magnesium reacts with Chlorophosphonazo III (CPZ III) and
causes an increase in absorbance. EGTA is added to inhibit calcium
interference. Addition of EDTA to the reaction allows for accurate sample
blanking. The reaction is a two-point, end-point method that is measured
photometrically at 660 nm.
In this method calcium reacts with 5-nitro-5’-methyl-BAPTA (NM-BAPTA) in an
alkaline environment to form a complex that is measured at 340nm. When EDTA is
added to the reaction, two new complexes form, and the change in absorbance is
directly proportional to the calcium concentration.
This method is an enzymatic process, based on the oxidation of oxalate by
oxalate oxidase, followed by measurement of hydrogen peroxide produced during
the reaction by a peroxidase-catalyzed reaction. The primary measuring
wavelength is 600 nm, and the secondary measuring wavelength is 700 nm. The
procedure is specific for oxalate.
7. Urea Nitrogen
This method utilizes a coupled enzyme reaction (urease, followed by glutamate
dehydrogenase), with measurement of NADH (converting to NAD+) occurring at 340
Refer to the Laboratory Method Files section for detailed laboratory
procedure manual(s) of the methods used.
Laboratory Method Files
Albumin - Kidney Markers - 24-Hour Urine - 2nd Collection
Creatinine - Kidney Markers - 24-Hour Urine - 2nd Collection
Assurance and Monitoring
Urine specimens were processed, stored, and shipped to University of
Minnesota, Minneapolis, MN for analysis. Detailed instructions on specimen
collection and processing are discussed in the NHANES
24-Hour Urine Study Procedures Manual. Vials are stored under appropriate
frozen (-30°C) conditions until they are shipped to University of Minnesota for
The NHANES quality assurance and quality control (QA/QC) protocols meet the
1988 Clinical Laboratory Improvement Act mandates. Detailed QA/QC instructions
are discussed in the NHANES Laboratory
Procedures Manual (LPM).
Mobile Examination Centers (MECs)
Laboratory team performance is monitored using several techniques. NCHS and
contract consultants use a structured quality assurance evaluation during
unscheduled visits to evaluate both the quality of the laboratory work and the
quality-control procedures. Each laboratory staff member is observed for
equipment operation, specimen collection and preparation; testing procedures and
constructive feedback are given to each staff member. Formal retraining sessions
are conducted annually to ensure that required skill levels were maintained.
NHANES uses several methods to monitor the quality of the analyses performed
by the contract laboratories. In the MEC, these methods include performing blind
split samples collected on “dry run” sessions. In addition, contract
laboratories randomly perform repeat testing on 2% of all specimens.
NCHS developed and distributed a quality control protocol for all the
contract laboratories which outlined the use of Westgard rules (Westgard et al.,
1981) when running NHANES specimens. Progress reports containing any problems
encountered during shipping or receipt of specimens, summary statistics for each
control pool, QC graphs, instrument calibration, reagents, and any special
considerations are submitted to NCHS quarterly. The reports are reviewed for
trends or shifts in the data. The laboratories are required to explain any
identified areas of concern.
Data Processing and Editing
The data were reviewed. Incomplete data or improbable values were sent to the
performing laboratory for confirmation.
Variables with data from 24-hour urine first collection and second collection
specimens are denoted with a “UR1” and “UR2” prefix, respectively.
Refer to the 2013
- 2014 Laboratory Data Overview for general information on NHANES laboratory
NHANES participants were selected on the basis of a national probability
design. In order to increase the number of participants for specific
demographic groups, a multi-stage, unequal probability of selection design was
implemented. In 2014, the sample design included an oversample of certain
demographic groups: persons 80 years and older, non-Hispanic black, non-Hispanic
Asian, Hispanic, and low income persons. Sample weights are constructed that
encompass the unequal probabilities of selection, as well as adjustments for
non-participation by selected sample persons. In order to produce national,
representative estimates, the appropriate sample weights must be used.
The 24-hour urine collection was conducted for a random one-half subsample of
1,103 persons 20-69 years examined in the mobile examination center in 2014. A
special one-year urine sample weight (WT1YUR) is provided for the 827
participants who provided a complete 24-hour urine specimen to obtain
representative national estimates based on this first 24-hour urine collection.
As with other NHANES subsamples, the 24-hour urine sample weights account for
the additional probability of selection into the subsample component as well as
the additional nonresponse. This sample weight, along with strata (VSTRA) and
PSUs (VPSU) are used to calculate variance estimates based on the Taylor Series
Linearization method. In addition, 16 Fay-adjusted balanced repeated replication
(Fay’s BRR) weights (WT1YUB01 – WT1YUB16), along with their corresponding strata
(VSTRABRR) and PSUs (VPSUBRR), and 15 Jackknife replicate weights (WT1YUR01 –
WT1YUR15) that can be used with VSTRA and VPSU are included in the files to
obtain variance estimates based on a replication method. The Fay’s BRR weights
were created with an adjustment factor of 0.3. There is no separate sample
weight provided for use with the data collected for the second 24-hour urine
specimen. These data were collected for the purpose of estimating within-person
variability and not for obtaining nationally representative estimates based on
the second 24-hour urine sample. New sample weights would need to be calculated
by the user if any additional analytical efforts were planned for the second
Demographic and Other Related Variables
The analysis of NHANES laboratory data must be conducted using the
appropriate survey design and demographic variables. The NHANES
2013-2014 Demographics File contains demographic data, health indicators,
and other related information collected during household interviews.
Please see Appendix 1 for the list of data files produced from the 24-hour
This laboratory data file can be linked to the other NHANES data files using
the unique survey participant identifier (i.e., SEQN).
The detection limits were constant for all of the analytes in the data set.
Two variables are provided for each of these analytes. The variable name ending
in “LC” (ex., UR1MALC) indicates whether the result was below the limit of
detection: the value “0” means that the result was at or above the limit of
detection, “1” indicates that the result was below the limit of detection.
For analytes with analytic results below the lower limit of detection (ex.,
UR1MALC=1), an imputed fill value was placed in the analyte results field. This
value is the lower limit of detection divided by the square root of 2
(LLOD/sqrt). The other variable prefixed UR1 (ex., UR1MA) provides the
analytic result for that analyte.
The lower limit of detection (LLOD) for 24-Hour Urine Kidney Markers First
and Second Collections:
||Albumin, Urine 1st Collection (µg/mL)
||Albumin, Urine 2nd Collection (µg/mL)
||Calcium, Urine 1st Collection (mg/dL)
||Calcium, Urine 2nd Collection (mg/dL)
||Creatinine, Urine 1st Collection (mg/dL)
||Creatinine, Urine 2nd Collection (mg/dL)
||Magnesium, Urine 1st Collection (mg/dL)
||Magnesium, Urine 2nd Collection (mg/dL)
||Oxalate, Urine 1st Collection (mg/L)
||Oxalate, Urine 2nd Collection (mg/L)
||Phosphorus, Urine 1st Collection (mg/dL)
||Phosphorus, Urine 2nd Collection (mg/dL)
||Urea Nitrogen, 1st Collection (mg/dL)
||Urea Nitrogen, 2nd Collection (mg/dL)
Please refer to the NHANES Analytic
Guidelines and the on-line NHANES Tutorial
for further details on the use of sample weights and other analytic