ACCESS

Immunoassay Systems
Access SHBG
Instructions For Use Sex Hormone-Binding Globulin
© 2021 Beckman Coulter, Inc. All rights reserved.
A48617

FOR PROFESSIONAL USE ONLY

PRINCIPLE
INTENDED USE

The Access SHBG assay is a paramagnetic particle, chemiluminescent immunoassay for the quantitative determination
of Sex Hormone Binding Globulin levels in human serum and plasma using the Access Immunoassay Systems.

SUMMARY AND EXPLANATION

Sex hormone-binding globulin (SHBG) is a glycoprotein responsible for blood transport of testosterone and estradiol.
SHBG is synthesized in the liver and has a high binding affinity for 17-hydroxysteroid hormones.1,2 Less than 2% of
biologically active steroids are free in the circulation with the remainder being bound mostly to SHBG and albumin.
SHBG has a high binding affinity to the 17-hydroxysteroid hormones while albumin has a low binding affinity.3 Initially,
the free portion or unbound hormone fraction was believed to be the only biologically active form. It is now recognized
that the portion of hormone that is weakly bound to albumin is also available to the tissues. The free hormone plus the
albumin bound portion of hormones represents the “bioavailable” hormone.4
The measurement of SHBG can be an important indicator of a chronic or excessive androgenic activity where clinical
symptoms would seem to indicate androgen in excess, but androgen levels are normal. Elevated SHBG levels can
be seen in persons with androgen insensitivities, hyperthyroidism, cirrhosis of the liver and is found in patients on oral
contraceptives or antiepileptic drugs.4,5,6 Decreased concentrations of SHBG are often seen in men with hypothyroidism
and androgen replacement therapy; where women with hirsutism, virilism, polycystic ovarian syndrome (PCOS), elevated
androgen levels, obesity and acromegaly will also see a decrease in SHBG levels.4,7,8
SHBG production is regulated by the androgen/estrogen balance, thyroid hormones, insulin, and dietary factors.4
The concentration of SHBG is increased by estrogens and decreased by androgens. Therefore, SHBG production
is stimulated by estradiol and suppressed by testosterone. As a result, SHBG concentrations are higher in women
versus men.9 Pregnant women have markedly higher SHBG serum concentrations due to their increased estrogen
production.8
When assessing a patient's androgen status for various conditions, physicians must consider using more than total
testosterone measurements. In addition to SHBG, “free” and “bioavailable” testosterone calculations can be used to
provide a better evaluation of androgen status.10,11
Free testosterone can be measured directly by equilibrium dialysis. Alternatively, the non-SHBG-bound fraction may
be obtained by precipitation of SHBG-bound testosterone with ammonium sulfate. As both methods are not routinely
performed in most laboratories, an indirect method calculation can be utilized to estimate free testosterone. Calculating
the Free Testosterone Index (FTI) or the Free Androgen Index (FAI) requires the measurement of total testosterone and
SHBG concentrations.12 The FAI is calculated using the equation:

The FAI is generally considered useful in estimating free testosterone in women with hirsutism or hyperandrogenism.

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Equations have been used to derive the free testosterone concentration (FTc). Vermeulen et al. described a
mathematical algorithm that incorporates the measurement of total testosterone and SHBG concentrations, that is
comparable to free testosterone values obtained by equilibrium dialysis.12 The Vermeulen equation is useful for
estimating free testosterone, except in pregnant patients.12
To calculate the “bioavailable” testosterone (BATc), the concentrations of SHBG, total testosterone (TT) and
albumin-bound testosterone must be considered. The calculated bioavailable testosterone is the sum of free
testosterone and albumin-bound testosterone.12 A detailed description of both the Vermeulen “free” and “bioavailable”
testosterone calculations can be found at [Link]

METHODOLOGY

The Access SHBG assay is a sequential two-step immunoenzymatic (“sandwich”) assay. A sample is added to a reaction
vessel along with paramagnetic particles coated with monoclonal anti-SHBG antibody and saline buffer with proteins.
After incubation in a reaction vessel, materials bound to the solid phase are held in a magnetic field while unbound
materials are washed away. A second monoclonal anti-SHBG antibody conjugated to alkaline phosphatase is added to
the reaction vessel.
After the second incubation in the reaction vessel, materials bound to the solid phase are held in a magnetic field while
unbound materials are washed away. Then, the chemiluminescent substrate is added to the vessel and light generated
by the reaction is measured with a luminometer. The light production is directly proportional to the concentration of
SHBG in the sample. The amount of analyte in the sample is determined from a stored, multi-point calibration curve.

SPECIMEN
SPECIMEN COLLECTION AND PREPARATION

1. Serum and plasma (Heparin) are the recommended samples.

2. Observe the following recommendations for handling, processing, and storing blood samples:13
• Collect all blood samples observing routine precautions for venipuncture.
• Allow serum samples to clot completely before centrifugation.
• Keep tubes stoppered at all times.
• Physically separate serum or plasma from contact with cells as soon as possible.
• Store samples tightly stoppered at room temperature (15 to 30°C) for no longer than eight hours.
• If the assay will not be completed within eight hours, refrigerate the samples at 2 to 8°C.
• For serum samples:
- If the assay will not be completed within 7 days, or for shipment of samples, freeze at -20°C or colder up to
2 months.
- Thaw samples no more than three times.
• For plasma (heparin) samples:
- If the assay will not be completed within 5 days, or for shipment of samples, freeze at -20°C or colder up to
2 months.
- Thaw samples no more than two times.
3. Use the following guidelines when preparing specimens:
• Ensure residual fibrin and cellular matter have been removed prior to analysis.
• Follow blood collection tube manufacturer's recommendations for centrifugation.

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 4. Each laboratory should determine the acceptability of its own blood collection tubes and serum separation products.
Variations in these products may exist between manufacturers and, at times, from lot-to-lot.
5. Avoid assaying lipemic or hemolyzed samples.

REAGENTS
PRODUCT INFORMATION

Access SHBG Reagent Pack

Cat. No. A48617: 100 determinations, 2 packs, 50 tests/pack
• Provided ready to use.
• Store upright and refrigerate at 2 to 10°C.
• Refrigerate at 2 to 10°C for a minimum of two hours before use on the instrument.
• Stable until the expiration date stated on the label when stored at 2 to 10°C.
• Stable at 2 to 10°C for 28 days after initial use.
• Signs of possible deterioration are a broken elastomeric layer on the pack or control values out of range.
• If the reagent pack is damaged (i.e., broken elastomer), discard the pack.

R1a: Paramagnetic particles coated with mouse monoclonal anti-SHBG, protein (bovine, mouse)
buffered matrix, < 0.1% sodium azide, 0.1% ProClin* 300.
R1b: Mouse monoclonal anti-SHBG alkaline phosphatase (bovine) conjugate, buffered matrix with
protein (bovine), < 0.1% sodium azide, 0.1% ProClin 300.
R1c: TRIS buffer with < 0.1% sodium azide and 0.1% ProClin 300.

*ProClin™ is a trademark of The Dow Chemical Company (“Dow”) or an affiliated company of Dow.

WARNING AND PRECAUTIONS

• For in vitro diagnostic use.

• Patient samples and blood-derived products may be routinely processed with minimum risk using the procedure
described. However, handle these products as potentially infectious according to universal precautions and good
clinical laboratory practices, regardless of their origin, treatment, or prior certification. Use an appropriate disinfectant
for decontamination. Store and dispose of these materials and their containers in accordance with local regulations
and guidelines.
• For hazards presented by the product refer to the following sections: REACTIVE INGREDIENTS and GHS HAZARD
CLASSIFICATION.

REACTIVE INGREDIENTS

CAUTION
Sodium azide preservative may form explosive compounds in metal drain lines.
See NIOSH Bulletin: Explosive Azide Hazard (8/16/76).
To avoid the possible build-up of azide compounds, flush wastepipes with
water after the disposal of undiluted reagent. Sodium azide disposal must be in
accordance with appropriate local regulations.

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GHS HAZARD CLASSIFICATION

SHBG PMP with blockers WARNING

(Compartment R1a)

H317 May cause an allergic skin reaction.

H412 Harmful to aquatic life with long lasting effects.
P273 Avoid release to the environment.
P280 Wear protective gloves, protective clothing and eye/face
protection.
P333+P313 If skin irritation or rash occurs: Get medical
advice/attention.
P362+P364 Take off contaminated clothing and wash it before use.
reaction mass of: 5-chloro-2-methyl-4-isothiazolin -3-one
[EC# 247-500-7] and 2-methyl-4-isothiazolin-3-one [EC#
220-239-6](3:1) < 0.05%
SHBG Conjugate WARNING
(Compartment R1b)

H317 May cause an allergic skin reaction.

H412 Harmful to aquatic life with long lasting effects.
P273 Avoid release to the environment.
P280 Wear protective gloves, protective clothing and eye/face
protection.
P333+P313 If skin irritation or rash occurs: Get medical
advice/attention.
P362+P364 Take off contaminated clothing and wash it before use.
reaction mass of: 5-chloro-2-methyl-4-isothiazolin -3-one
[EC# 247-500-7] and 2-methyl-4-isothiazolin-3-one [EC#
220-239-6](3:1) < 0.05%
SHBG Ancillary (Compartment WARNING
R1c)

H317 May cause an allergic skin reaction.

H412 Harmful to aquatic life with long lasting effects.
P273 Avoid release to the environment.

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 P280 Wear protective gloves, protective clothing and eye/face
protection.
P333+P313 If skin irritation or rash occurs: Get medical
advice/attention.
P362+P364 Take off contaminated clothing and wash it before use.
reaction mass of: 5-chloro-2-methyl-4-isothiazolin -3-one
[EC# 247-500-7] and 2-methyl-4-isothiazolin-3-one [EC#
220-239-6](3:1) < 0.05%

Safety Data Sheet is available at [Link]/techdocs

MATERIALS NEEDED BUT NOT SUPPLIED WITH REAGENT KIT

1. Access SHBG Calibrators

Provided at zero and approximately 3, 9, 27, 80 and 200 nmol/L (approximately 3, 9, 27, 80 and 200 IU/mL).
Cat. No. A48618
2. Access SHBG Quality Control (QC)
Provided at approximately 10 and 100 nmol/L (approximately 10 and 100 IU/mL).
Cat. No. A48619
3. Access Substrate
Cat. No. 81906
4. Access Wash Buffer II, Cat. No. A16792
UniCel DxI Wash Buffer II, Cat. No. A16793
UniCel DxI Access Immunoassay Systems Wash Buffer II, Cat. No A79784
(Diluent pack for use with the UniCel DxI system onboard dilution feature.)

EQUIPMENT AND MATERIALS

R1 Access SHBG Reagent Packs

CALIBRATION
CALIBRATION INFORMATION

An active calibration curve is required for all tests. For the Access SHBG assay, calibration is required every 28 days.
Refer to the appropriate system manuals and/or Help system for information on calibration theory, configuring calibrators,
calibrator test request entry, and reviewing calibration data.

QUALITY CONTROL
Quality control materials simulate the characteristics of patient samples and are essential for monitoring the system
performance of immunochemical assays. Because samples can be processed at any time in a “random access” format
rather than a “batch” format, quality control materials should be included in each 24-hour time period.14 Include Access
SHBG QC or other commercially available quality control materials that cover at least two levels of analyte. More frequent
use of controls or the use of additional controls is left to the discretion of the user based on good laboratory practices
or laboratory accreditation requirements and applicable laws. Follow manufacturer's instructions for reconstitution and
storage. Each laboratory should establish mean values and acceptable ranges to assure proper performance. Quality

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control results that do not fall within acceptable ranges may indicate invalid test results. Examine all test results generated
since obtaining the last acceptable quality control test point for this analyte. Refer to the appropriate system manuals
and/or Help system for information about reviewing quality control results.
The Access SHBG assay has been evaluated at an ambient temperature range of 18-32°C. Should your ambient
laboratory temperature fluctuate > ± 5°C within this range, carefully review the daily quality control results and
recalibrate as necessary.

TESTING PROCEDURE(S)
PROCEDURAL COMMENTS
1. Refer to the appropriate system manuals and/or Help system for a specific description of installation, start-up,
principles of operation, system performance characteristics, operating instructions, calibration procedures,
operational limitations and precautions, hazards, maintenance, and troubleshooting.
2. Mix contents of new (unpunctured) reagent packs by gently inverting pack several times before loading on the
instrument. Do not invert open (punctured) packs.
3. Use twenty (20) µL of sample for each determination in addition to the sample container and system dead volumes.
Use fifty (50) µL of sample in addition to the sample container and system dead volumes for each determination
run with the DxI system onboard dilution feature. Refer to the appropriate system manuals and/or Help system for
the minimum sample volume required.
4. The system default unit of measure for sample results is nmol/L. To change sample reporting units to the
International System of Units (SI units), refer to the appropriate system manuals and/or Help system. To
manually convert concentrations to the International System, 1 nmol/L is equal to 1 IU/mL. To manually convert
concentrations to µg/mL, multiply nmol/L by multiplication factor 0.095.

PROCEDURE
Refer to the appropriate system manuals and/or Help system for information on managing samples, configuring tests,
requesting tests, and reviewing test results.

RESULTS INTERPRETATION
Patient test results are determined automatically by the system software. The amount of analyte in the sample is
determined from the measured light production by means of the stored calibration data. Patient test results can be
reviewed using the appropriate screen. Refer to the appropriate system manuals and/or Help system for complete
instructions on reviewing sample results.

REPORTING RESULTS
EXPECTED RESULTS

1. Each laboratory should establish its own reference ranges to assure proper representation of specific populations.
2. SHBG production is regulated by the androgen/estrogen balance, thyroid hormones, insulin and dietary factors.1
The concentration of SHBG is increased by estrogens and decreased by androgens. Therefore, SHBG is stimulated
by estradiol and suppressed by testosterone. As a result, SHBG concentrations are higher in women versus men
and contraceptive treatment may increase SHBG concentrations. Pregnant women also have markedly higher
SHBG serum concentrations due to their increased estrogen production.
3. For the expected values presented below, samples were selected from an apparently healthy population.
Apparently healthy is defined as general health and appearance, considered to be normal with no known or
apparent diseases, or medications that would influence Access SHBG results.
• Male (20-50 years) samples included in the study had no known pre-existing endocrine disorders with Access
Testosterone values > 1.75 ng/mL, and Access hTSH values between 0.34-5.60 µIU/mL.

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 • Female (20-46 years) samples included in the study were non-pregnant, normal cycling women not using any
steroid based contraceptives, with no known endocrine or reproductive disorders. Additionally, samples with
Access Total βhCG values < 2.9 mIU/mL, Access Estradiol values > 20 pg/mL, and Access hTSH values between
0.34-5.60 µIU/mL were included.
• Postmenopausal female (47-91 years) samples included in the study were non-menstruating for 12 or more
consecutive months, not on hormone replacement therapy, and not taking any osteoporosis medication with
estrogen analog.
For women < 55 years, confirmation of menopause was determined by Access hLH, Access hFSH
measurements consistent with postmenopausal status, and Access Estradiol measurements < 88 pg/mL.
Additionally, samples with Access hTSH values between 0.34-5.60 µIU/mL were included.
4. Median values are 95% non parametric reference interval populations are described below. A detailed
description about the calculation procedures are available on request. Refer also to the homepage of
[Link]/[Link].
5. Reference Interval data provided in the tables below were obtained with Access SHBG and Access Testosterone
assays on a UniCel DxI 800. If other assays are used, the user should determine reference intervals for the specific
assays.
6. All index calculations should be performed with results generated using Access SHBG and Access Testosterone
assays.

Table 1.0 SHBG Reference Interval Data:

Median Value 95% Confidence
SHBG n Median Age (nmol/L) Interval (nmol/L)
Males 20-50 years 151 37 38.2 13.3-89.5
Females 20-46 years 141 34 47.2 18.2-135.5
Females 47-91 years 131 59 49.6 16.8-125.2
(post-menopausal)

Free Androgen Index (FAI):

The FAI is an indirect method used to assess androgen levels.
• Note: Use the same units for total testosterone (TT) and SHBG when Calculating the FAI%
• The FAI is a ratio of results for total testosterone (TT) and SHBG.

Table 2.0 FAI Reference Interval Data:

Free Androgen 95% Confidence
Index (%) n Median Age Median Value (%) Interval (%)
Males 20-50 years 151 37 51.6 24.3-110.2
Females 20-46 years 141 34 2.67 0.65-10.93
Females 47-91 years 131 59 1.70 0.23-6.80
(post-menopausal)

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Free Testosterone (FT):
This is an indirect method used to estimate free circulating testosterone. This value may be calculated from results
generated using Access SHBG, Access Testosterone and albumin concentrations.
• The FT concentration may be calculated as Apparent Free Testosterone Concentration (AFTC).
• Calculations can be completed at the homepage of [Link]/[Link].

Table 3.0 AFTC Reference Interval Data:

Median Value 95% Confidence
AFTC (nmol/L) n Median Age (nmol/L) Interval (nmol/L)
Males 20-50 years 151 37 0.38 0.17-0.66
Females 20-46 years 141 34 0.019 0.006-0.055
Females 47-91 years 131 59 0.011 0.002-0.033
(post-menopausal)

Bioavailable Testosterone:
The sum of albumin-bound testosterone plus free testosterone provides the bioavailable testosterone value. This value
may be calculated using total testosterone, SHBG and albumin concentrations.
• The reported information can be presented in either units (nmol/L) or percentage.
• A detailed description about the calculation procedures are available on request.
• Calculations can be completed at the homepage of [Link]/[Link].

Table 4.0 Bioavailable Testosterone Reference Intervals:

Bioavailable
Testosterone Median Value 95% Confidence
(nmol/L) n Median Age (nmol/L) Interval (nmol/L)
Males 20-50 years 151 37 9.0 4.0-15.5
Females 20-46 years 141 34 0.44 0.13-1.30
Females 47-91 years 131 59 0.265 0.043-0.763
(post-menopausal)

Table 5.0 Bioavailable Testosterone Percentage Reference Intervals:

Bioavailable 95% Confidence
Testosterone (%) n Median Age Median Value (%) Interval (%)
Males 20-50 years 151 37 46.8 25.7-70.2
Females 20-46 years 141 34 33.6 14.8-57.4
Females 47-91 years 131 59 31.0 13.3-59.0
(post-menopausal)

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PROCEDURAL NOTES
LIMITATIONS

1. Samples can be accurately measured within the analytical range of the lower limit of detection (0.33 nmol/L) and
the highest calibrator value (approximately 200 nmol/L).
• If a sample contains less than the lower limit of detection for the assay, report the results as less than that value
(i.e., < 0.33 nmol/L, < 0.03 µg/mL). When the DxI system onboard dilution feature is used, the system will report
results as less than 170 nmol/L (16.15 µg/mL).
• If a sample contains more than the stated value of the highest Access SHBG Calibrator (S5), report the result
as greater than the value of the highest calibrator. Alternatively, dilute one volume of sample with 9 volumes of
Wash Buffer II.
• Refer to the appropriate system manuals and/or Help system for instructions on entering a sample dilution in a
test request. The system reports the results adjusted for the dilution.
The DxI system onboard dilution feature automates the dilution process, using one volume of sample with nine
volumes of UniCel DxI Access Immunoassay Systems Wash Buffer II, allowing samples to be quantitated up to
approximately 2,000 nmol/L (190 µg/mL). The system reports the results adjusted for the dilution.
2. For assays employing antibodies, the possibility exists for interference by heterophile antibodies in the patient
sample. Patients who have been regularly exposed to animals or have received immunotherapy or diagnostic
procedures utilizing immunoglobulins or immunoglobulin fragments may produce antibodies, e.g. HAMA, that
interfere with immunoassays. Additionally, other heterophile antibodies such as human anti-goat antibodies may
be present in patient samples.15,16
Such interfering antibodies may cause erroneous results. Carefully evaluate the results of patients suspected of
having these antibodies.
3. For patients presenting with cirrhosis6 or sub-clinical thyroid conditions,17,18 carefully evaluate results as this
condition can potentially cause erroneously elevated SHBG results.
4. The Access SHBG results should be interpreted in light of the total clinical presentation of the patient, including:
symptoms, clinical history, data from additional tests, and other appropriate information.
5. The Access SHBG assay does not demonstrate any “hook” effect up to 49,500 nmol/L.

PERFORMANCE CHARACTERISTICS
PERFORMANCE CHARACTERISTICS

METHODS COMPARISON
A comparison of 158 values using the Access SHBG assay on the UniCel DxI 800 immunoassay system and
a commercially available enzyme immunoassay system gave the following statistical data using non-parametric
Passing-Bablok regression calculations:

Intercept Slope
Range of (95% Confidence (95% Confidence Correlation
n Observations (nmol/L) Interval) (nmol/L) Interval) Coefficient (r2)
158 5.7-184.5 1.84 1.09 0.94
(0.54-3.00) (1.06-1.12)

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Table 6.0 Demographics
Age Range
Subject Category Number of Subjects Gender Age Mean (years) (years)
Pregnancy 34 34 F 30 20-45
Liver Disease 24 20 M - 4 F 54 21-77
PCOS 18 18 F 32 18-92
Hyperthyroid 8 2M-6F 63 32-78
Hypogonadism 15 15 M 45 16-69
Hypothyroid 7 7 F 61 34-77
Oral Contraceptive 4 4 F 29 24-39
Sterility 5 5 F 28 19-34
Diabetes 4 2M-2F 56 53-63
Other 39 11 M - 28 F 47 9-93

DILUTION RECOVERY (LINEARITY)

Samples can be accurately measured within the analytical range of the lower limit of detection and the highest calibrator
value (0.33-200 nmol/L). Data was generated using a UniCel DxI 800.

Expected Concentration Determined Concentration

Sample 1 (nmol/L) (nmol/L) Recovery (%)
Neat 206.58 206.58 N/A
1:1.2 172.08 173.16 101
1:1.5 137.79 143.94 104
1:2 103.29 105.66 102
1:3 68.79 70.84 103
1:10 20.66 21.36 103
Mean % Recovery 103

Expected Concentration Determined Concentration

Sample 2 (nmol/L) (nmol/L) Recovery (%)
Neat 35.81 35.81 N/A
1:1.2 29.83 31.65 106
1:1.5 23.89 24.93 104
1:2 17.91 17.69 99
1:3 11.93 11.88 100
1:10 3.59 3.46 96
Mean % Recovery 101

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 Expected Concentration Determined Concentration
Sample 3 (nmol/L) (nmol/L) Recovery (%)
Neat 2.90 2.90 N/A
1:1.2 2.42 2.41 100
1:1.5 1.94 1.96 101
1:2 1.45 1.48 102
1:3 0.97 0.99 102
1:10 0.29 0.28 96
Mean % Recovery 100

SPIKING RECOVERY
The addition of different levels of SHBG to three patient samples, on the UniCel DxI 800, resulted in the following data:

Expected Concentration Determined Concentration

Sample 1 (nmol/L) (nmol/L) Recovery (%)
Neat N/A 14.7 N/A
Low 46 50 108
Low-medium 78 77 99
Medium 110 110 100
Medium-high 141 143 101
High 173 174 100
High-end 217 205 94
Mean % Recovery 101

Expected Concentration Determined Concentration

Sample 2 (nmol/L) (nmol/L) Recovery (%)
Neat N/A 14.3 N/A
Low 46 44 95
Low-medium 78 78 101
Medium 109 110 101
Medium-high 141 137 97
High 173 172 100
High-end 217 199 92
Mean % Recovery 98

Expected Concentration Determined Concentration

Sample 3 (nmol/L) (nmol/L) Recovery (%)
Neat N/A 14.7 N/A
Low 46 46 98

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 Expected Concentration Determined Concentration
Sample 3 (nmol/L) (nmol/L) Recovery (%)
Low-medium 78 79 101
Medium 110 113 103
Medium-high 141 146 103
High 170 164 96
High-end 217 201 92
Mean % Recovery 99

IMPRECISION
This assay exhibits total imprecision of < 7% at concentrations greater than 2 nmol/L. The study, which consisted of
four patient samples at varying SHBG levels, on three separate pack lots, in duplicate, running for 20 different days,
completing 2 runs per day, over a period of 28 days on two UniCel DxI 800 instruments, provided the following data,
analyzed via analysis of variance (ANOVA).19

Mean (n = 480) Within-run Total Imprecision

Sample (nmol/L) SD CV % SD CV %
1 6.3 0.3 4.7 0.3 5.4
2 38 1.8 4.6 2.0 5.3
3 80 3.6 4.5 4.4 5.5
4 171 8.1 4.8 9.0 5.2
A polynomial regression of all imprecision data (4 samples spanning the range of the assay), generated from 3 reagent
lots and analyzed on multiple instruments, provides an overall estimation of the imprecision. Data was further analyzed
utilizing linear modeling plotting the log of the sample concentration by the log of the standard deviation, as shown in the
following table, and determining the upper limit of 95% confidence interval of this regression fit.

Dose (nmol/L) Estimated CV % Estimated SD

2 5.3 0.11
5 5.3 0.27
10 5.3 0.53
15 5.3 0.79
25 5.3 1.32
40 5.3 2.11
60 5.3 3.17
80 5.3 4.22
100 5.3 5.27
150 5.3 7.89
180 5.3 9.47

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ANALYTICAL SPECIFICITY / INTERFERENCES
The following interferents were added to low and high human serum samples, using the UniCel DxI 800, at the
concentrations indicated in the table below. All SHBG values obtained in the presence of each interferent indicate that
these substances do not significantly interfere with the assay, per CLSI EP7-A2.20
Note: ND = interference not detectable.

Substance Added Concentration Added Expected (nM) Observed (nM) % Interference

Low High Low High Low High
Acetamin- 30 mg/L 12.66 143.98 12.90 147.74 ND 2.6
ophen
200 mg/L 12.66 143.98 13.08 145.72 3.4 ND
Acetylsali- 390 mg/L 13.47 145.65 12.73 142.19 ND -2.4
cylic
acid
652 mg/L 13.47 145.65 12.60 146.25 -6.4 ND
Alpha-Feto- 74.9 µg/L 14.26 153.83 14.62 160.71 2.6 4.5
protein
(AFP)
492 µg/L 14.26 153.83 14.49 153.74 ND ND
Bilirubin, conjugated 2.98 mg/L 12.84 139.81 12.88 140.12 ND ND
300 mg/L 12.84 139.81 12.82 146.08 ND 4.5
Bilirubin, unconjugated 12.2 mg/L 13.14 138.88 12.19 142.15 -7.3 2.4
200 mg/L 13.14 138.88 12.45 146.16 -5.3 5.2
Cortisol 16 µg/L 12.39 139.05 12.04 138.56 -2.8 ND
100 mg/L 12.39 139.05 12.50 137.88 ND ND
11-deoxy- 1.58 mg/L 12.17 136.97 12.16 139.46 ND ND
cortisol
4 mg/L 12.17 136.97 12.45 139.45 ND ND
5α-dihydroxy- 0.85 µg/L 11.92 136.67 12.06 138.76 ND ND
testosterone
20 mg/L 11.92 136.67 12.96 148.96 8.8 9.0
Hemoglobin 2 g/L 11.10 126.19 11.35 122.28 ND -3.1
4 g/L 11.10 126.19 11.66 127.64 5.0 ND
Heparin 1,000 U/L 13.44 147.43 13.98 152.09 4.1 3.2
72,000 U/L 13.44 147.43 13.38 152.18 ND 3.2
Human serum albumin 51 g/L 13.02 143.62 12.62 137.06 ND -4.6
(HSA)
60 g/L 13.02 143.62 12.60 138.50 ND -3.6
Ibuprofen 0.07 g/L 13.47 146.45 13.28 147.63 ND ND
0.5 g/L 13.46 145.45 12.95 147.67 -3.8 ND
Estradiol 750 ng/L 12.81 140.64 12.44 144.82 -2.9 3.0

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 Substance Added Concentration Added Expected (nM) Observed (nM) % Interference
Low High Low High Low High
3.6 µg/L 12.81 140.64 12.37 139.81 -3.4 ND
GAS6 63 µg/L 13.75 152.73 13.51 153.57 ND ND
235 µg/L 13.75 152.73 13.21 151.28 -3.9 ND
Laminin 520 µg/L 11.95 137.38 12.26 137.29 ND ND
6,064 µg/L 11.95 137.38 12.07 139.25 ND ND
Multivitamin 0.3% (v/v) 13.22 149.27 13.12 148.59 ND ND
supplement
0.9% (v/v) 13.22 149.27 13.56 151.36 ND ND
Protein S 3.6 mg/L 11.98 142.20 12.28 137.92 2.5 -3.0
26 mg/L 11.98 142.20 11.98 138.82 ND ND
Testosterone 210 ng/L 11.63 130.05 11.33 127.88 ND ND
20 mg/L 11.63 130.05 12.69 134.63 9.1 3.5
Thyroglobulin (Tg) 40 µg/L 14.01 152.32 13.85 152.61 ND ND
300 µg/L 14.01 152.32 14.18 152.60 ND ND
Thyroxine- 55 mg/L 11.64 126.75 12.13 129.16 4.2 ND
binding Globulin (TBG)
193 mg/L 11.64 126.75 11.32 126.32 ND ND
Transferrin 1.3 g/L 12.05 138.00 12.14 134.55 ND ND
4 g/L 12.05 138.00 12.31 136.27 ND ND
Triglycerides (intralipid) 3.29 g/L 13.28 145.39 13.69 143.33 3.1 ND
50 g/L 9.71 111.14 10.13 118.34 4.4 6.5
To evaluate potential cross-reactivity of various molecules, the substances shown in the following table were spiked in
the Access SHBG Calibrator S0 and run on the UniCel DxI 800. Values for cross reactivity were calculated as described
in CLSI EP7-A2.20 No significant cross-reactivity was observed.
Note: ND = no detectable cross-reactivity.

Substance Added Concentration Added Concentration Observed % Cross Reactivity

Alpha-Fetoprotein (AFP) 74.9 µg/L ND ND
492 µg/L ND ND
Cortisol 16 µg/L ND ND
100 mg/L ND ND
5α-dihydroxytestosterone 0.85 µg/L ND ND
20 mg/L ND ND
11-deoxycortisol 1.58 mg/L ND ND
4 mg/L ND ND
Estradiol 750 ng/L ND ND
3.6 µg/L ND ND

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 Substance Added Concentration Added Concentration Observed % Cross Reactivity
GAS6 63 µg/L ND ND
235 µg/L ND ND
Laminin 520 µg/L ND ND
6,064 µg/L ND ND
Protein S 3.6 mg/L ND ND
26 mg/L ND ND
Testosterone 210 ng/L ND ND
20 mg/L ND ND
Thyroglobulin (Tg) 40 µg/L ND ND
300 µg/L ND ND
Thyroxine-binding Globulin 55 mg/L ND ND
(TBG)
193 mg/L ND ND
Transferrin 1.3 g/L ND ND
4 g/L ND ND

ANALYTICAL SENSITIVITY

Limit of Blank (Analytical Sensitivity)

Limit of Blank (LoB) for Access SHBG was determined to be 0.017 nmol/L. LoB was tested using a protocol based
on CLSI EP17-A.21 A total number of 156 replicates of a zero analyte sample (Access SHBG Calibrator S0) were
measured in 12 runs using three UniCel DxI 800 analyzers. The 95th percentile of the 156 replicates was estimated
using a non-parametric approach. The 97.5% upper confidence limit of this estimate was determined as LoB.

Limit of Detection
Limit of Detection (LoD) of Access SHBG assay was determined to be 0.33 nmol/L, based on the lowest level sample
where the beta-percentile (defined as the percentage of observations below LoB) was 5% or less. The LoD study was
run under a protocol based on CLSI EP17-A21 resulting in 36 replicates for each of five low-level samples (180 total)
measured in 12 runs on three UniCel DxI 800 analyzers.

ADDITIONAL INFORMATION
Beckman Coulter, the stylized logo, and the Beckman Coulter product and service marks mentioned herein are
trademarks or registered trademarks of Beckman Coulter, Inc. in the United States and other countries.
May be covered by one or more pat. -see [Link]/patents.

REVISION HISTORY

Revision M
IFU updated to add Dutch, Finnish, Macedonian, Traditional Chinese, and Estonian

Revision N
Remove CE Mark and EC REP Address, Update GHS Hazard Information, and Website Address

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SYMBOLS KEY

Glossary of Symbols is available at [Link]/techdocs (document number C02724).

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REFERENCES
1. Selby C. Sex hormone binding globulin: origin, function and clinical significance. Ann Clin Biochem 1990; 27:
532-541.

2. Munell F, Suarez-Quian C, Selva D, Tirado O, Reventos J. Androgen binding protein and reproduction: where do
we stand? J of Andrology 2002; 23: 598-609.

3. Manni A, Pardridge W, Cefalu W, Nisula B, Bardin CW, Santner S, Santen R. Bioavailability of albumin-bound
testosterone. J Clin Endocrinology and Metabolism 1985; 61: 705-710.

4. Burtis CA, Ashwood ER, Bruns DE. Tietz textbook of clinical chemistry and molecular diagnostics. Saunders, 2006.
p. 2011-2012.

5. Ford HC, Cooke RR, Keightley EA, Feek CM. Serum levels of free and bound testosterone in hyperthyroidism.
Clinical Endocrinology 1992; 36: 187-192.

6. Luppa PB, Thaler M, Schulte-Frohlinde E, Schreiegg A, Huber U, Metzger J. Unchanged androgen-binding

properties of sex hormone-binding globulin in male patients with liver cirrhosis. Clin Chem Lab Med 2006; 44(8):
967-973.

7. Kacsoh B. Endocrine physiology 4. McGraw-Hill, New York, NY, 2000; p. 49.

8. Belgorsky A, Escobar ME, Rivarola MA. Validity of the calculation of non-sex hormone-binding globulin-bound
estradiol from total testosterone, total estradiol and sex hormone-binding globulin concentrations in human serum.
J. steroid Biochem. 1987; 28: 429-432.

9. Elmlinger M, Kuhnel W, Wormstall H, Doller P. Reference intervals of testosterone, androstenedione and SHBG
levels in healthy females and males from birth to old age. Clin Lab 2005; 51 (11-12): 625-632.

10. Androgen Deficiency Syndromes in Men Guideline Task Force. Testosterone therapy in adult men with
androgen deficiency syndromes: an endocrine society clinical practice guideline. J Clin Endocrinol Metab
2006;91(6):1995-2010.

11. Martin KA, Chang RJ, Ehrmann DA, et al. Evaluation and treatment of hirsutism in premenopausal women: an
endocrine society clinical practice guideline. J Clin Endocrinol Metab 2008;93(4):1105-1120.

12. Vermeulen A, Verdonck L, Kaufman J, A critical evaluation of simple methods for the estimation of free testosterone
in serum. Journal of Clinical Endocrinology & Metabolism 1999; 84: 3666-3672.

13. Approved Guideline - Procedures for the Handling and Processing of Blood Specimens for Common Laboratory
Tests, GP44-A4. 2010. Clinical and Laboratory Standards Institute.

14. Cembrowski GS, Carey RN. Laboratory quality management: QC ⇌ QA. ASCP Press, Chicago, IL, 1989.

15. Kricka, L. Interferences in immunoassays - still a threat. Clin Chem 2000; 46: 1037-1038.

16. Bjerner J, et al. Immunometric assay interference: incidence and prevention. Clin Chem 2002; 48: 613-621.

17. Saller B, Broda N, Heydarian R, Görges R, Mann K. Utility of third generation thyrotropin assays in thyroid function
testing. Exp Clin Endocrinol Diabetes 1998; 106 (Suppl 4): S29-S33.

18. Beckett, GJ. The investigation of thyroid function. J Intl Fed Clin Chem 1994; 6 (5):186-190.

19. Approved Guideline - Evaluation of Precision Performance of Quantitative Measurement Methods, EP5-A2.
August 2004. Clinical and Laboratory Standards Institute.

Instructions For Use A87697 N English Access SHBG

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20. Approved Guideline - Interference Testing in Clinical Chemistry, EP7-A2. November 2005. Clinical and Laboratory
Standards Institute.

21. Approved Guideline - Protocols for Determination of Limits of Detection and Limits of Quantitation, EP17-A.
October 2004. Clinical and Laboratory Standards Institute.

Beckman Coulter, Inc., 250 S. Kraemer Blvd., Brea, CA 92821 U.S.A.

[Link]

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