HCG ER METHOD FOR THE TECHNICON IMMUNO 1 SYSTEM IN-VITRO DIAGNOSTIC CLINICAL METHOD

K961351 · Bayer Corp. · DHA · Jun 3, 1996 · Clinical Chemistry

Device Facts

• Record ID: K961351
• Device Name: HCG ER METHOD FOR THE TECHNICON IMMUNO 1 SYSTEM IN-VITRO DIAGNOSTIC CLINICAL METHOD
• Applicant: Bayer Corp.
• Product Code: DHA · Clinical Chemistry
• Decision Date: Jun 3, 1996
• Decision: SESE
• Submission Type: Traditional
• Regulation: 21 CFR 862.1155
• Device Class: Class 2

Intended Use

This in vitro diagnostic procedure is intended to quantitatively measure human chorionic gonadotropin (hCG) in human serum on the Technicon Immuno 1 system. Measurements of human chorionic gonadotropin are used for the detection of pregnancy. This method is an adaptation of an existing Technicon Immuno 1 hCG method and has an extended range from 800 mIU/mL to 100,000 mIU/mL. This diagnostic method is not intended for use on any other system.

Device Story

The hCG Extended Range (hCG ER) method is an adaptation of the standard Technicon Immuno 1 hCG assay, designed to measure high concentrations of hCG in human serum. The device uses a double antibody immunoassay principle; patient samples are reacted with hCG antibody conjugates and monoclonal ImmunoMagnetic Particles (mIMP) at 37°C. After washing, pNPP substrate is added; alkaline phosphatase in the conjugate converts pNPP to para-nitrophenoxide. The system monitors absorbance at 405nm and 450nm, where absorbance is directly proportional to hCG concentration. The ER method uses a smaller sample volume (1.3µL vs 50µL) and a multiplication factor of 50 to extend the reportable range to 100,000 mIU/mL. It is used in clinical laboratory settings on the Technicon Immuno 1 system. Healthcare providers use the quantitative results to detect pregnancy, particularly in cases where high hCG levels are expected, eliminating the need for manual sample dilution.

Clinical Evidence

Bench testing only. Precision evaluated over 20 days (n=130) with total CVs ranging from 4.8% to 6.8% across concentrations of 978 to 83,756 mIU/mL. Correlation study (n=104) comparing ER method to standard method yielded a regression equation of Y = 1.11X + 323 with r=0.998. Linearity and parallelism studies confirmed recovery within 95%-107% of expected values. No clinical patient outcome data provided.

Technological Characteristics

Double antibody immunoassay; uses monoclonal ImmunoMagnetic Particles (mIMP) and alkaline phosphatase-labeled antibody conjugates. Detection via spectrophotometry at 405nm and 450nm. Operates on the Technicon Immuno 1 system. Reagents and calibrators are identical to the standard hCG method; modification is limited to sample volume (1.3µL) and automated result calculation (multiplication factor of 50).

Indications for Use

Indicated for the quantitative measurement of human chorionic gonadotropin (hCG) in human serum for the detection of pregnancy in patients requiring an extended measurement range of 800 to 100,000 mIU/mL.

Regulatory Classification

Identification

A human chorionic gonadotropin (HCG) test system is a device intended for the early detection of pregnancy is intended to measure HCG, a placental hormone, in plasma or urine. A human chorionic goadotropin (HCG) test system is a device intended for any uses other than early detection of pregnancy (such as an aid in the diagnosis, prognosis, and management of treatment of persons with certain tumors or carcinomas) is intended to measure HCG, a placental hormone, in plasma or urine.

Predicate Devices

• Technicon Immuno 1 hCG method

Submission Summary (Full Text)

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K961351
Attachment 3
JUN - 3 1996

# Summary of Safety and Effectiveness

## hCG Extended Range (hCG ER) Method for the Technicon Immuno 1° system

Listed below is a comparison of the performance of the Technicon Immuno 1° hCG ER method and a similar method granted clearance of substantial equivalence (standard Technicon Immuno 1° hCG method). The information below was extracted from the Technicon Immuno 1 hCG ER method sheet, the Technicon Immuno 1 hCG method sheet, and data on file at Bayer Corporation.

## Intended Use

This *in vitro* diagnostic procedure is intended to quantitatively measure human chorionic gonadotropin (hCG) in human serum on the Technicon Immuno 1° system. Measurements of human chorionic gonadotropin are used for the detection of pregnancy. This method is an adaptation of an existing Technicon Immuno 1 hCG method and has an extended range from 800 mIU/mL to 100,000 mIU/mL. This diagnostic method is not intended for use on any other system.

| Part Number | Technicon
Immuno 1°
hCG ER
T01-2966-51 | | Technicon
Immuno 1°
hCG
T01-2966-51 | |
| --- | --- | --- | --- | --- |
| Reference Range | 800 - 100,000 mIU/mL | | 0.5 - 1000 mIU/mL | |
| Precision, within-run
(n = 130 over 20 days) | mean
(mIU/mL) | %CV | mean
(mIU/mL) | %CV |
| | 978 | 3.8% | 18.3 | 2.4% |
| | 9,707 | 3.5% | 55.7 | 2.2% |
| | 26,746 | 3.6% | 195.8 | 2.1% |
| | 28,658 | 3.8% | | |
| | 83,756 | 3.2% | | |
| Precision, total
(n = 130 over 20 days) | mean
(mIU/mL) | %CV | mean
(mIU/mL) | %CV |
| | 978 | 5.5% | 18.3 | 4.0% |
| | 9,707 | 6.2% | 55.7 | 3.7% |
| | 26,746 | 4.8% | 195.8 | 3.7% |
| | 28,658 | 6.8% | | |
| | 83,756 | 6.2% | | |

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Regression Equation

$$
Y = 1.11 X + 323
$$

where $Y = \text{Immuno 1 hCG ER}$

$X = \text{Immuno 1 hCG}$

$n = 104$ sera

$r = 0.998$

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# Assay Description

This method is a double antibody immunoassay. hCG Antibody Conjugate 1 (R1) and hCG Antibody Conjugate 2 (R2) are reacted with patient sample (or calibrator containing hCG) and incubated on the Technicon Immuno 1 system at $37^{\circ}\mathrm{C}$. The mIMP Reagent (monoclonal ImmunoMagnetic Particle) is added and a second incubation occurs during which the antibody complex is bound. The mIMP/antibody complex is then washed and the pNPP (para nitrophenylphosphate) substrate is added. The alkaline phosphatase (ALP) in the antibody conjugate reacts with the pNPP to form para-nitrophenoxide and phosphate. Increasing absorbance, due to the formation of para-nitrophenoxide, is monitored at $405\mathrm{nm}$ and $450\mathrm{nm}$. A sample having no hCG will have the minimum label bound, while samples containing high hCG concentrations will have maximum label bound. Thus the dose/response curve is directly proportional to the hCG in the sample.

The hCG ER method is an adaptation of the existing Technicon Immuno 1 hCG method. They both use the identical hCG Reagent, Part No. T01-2966-51. The hCG ER Calibrators, Part No. T03-3668-01, are identical in formulation to the Technicon SETpoint Reproductive Calibrators, Part No. T03-3148-01, which are used for the standard hCG method. There are no changes in either reagent or calibrator formulation. The hCG ER method differs from the standard hCG method in three ways:

1. The hCG ER calibrators, are reconstituted in $1.0\mathrm{mL}$ of Reproductive Calibrator Diluent while the Reproductive Calibrators are reconstituted in $2.0\mathrm{mL}$ of Reproductive Calibrator Diluent. This doubles the concentrations of hCG in the calibrator from 0 - 1000 mIU/mL to 0 - 2000 mIU/mL.

2. $65\mu \mathrm{L}$ of calibrator are reacted with reagent, while $1.3\mu \mathrm{L}$ of any other sample, serum or control, are reacted. $65\mu \mathrm{L}$ of the highest calibrator, $2000\mathrm{mIU / mL}$, corresponds to 130 total mIU of hCG. A corresponding serum sample with 130 total mIU of hCG, at $1.3\mu \mathrm{L}$ volume, would have $100,000\mathrm{mIU / mL}$ concentration. Thus a serum sample at $100,000\mathrm{mIU / mL}$ reacts the same total quantity of hCG, and produces the same amount of p-nitrophenoxide, as does calibrator at $2,000\mathrm{mIU / mL}$.

3. The concentrations of all samples other than calibrator are automatically multiplied by a factor of 50, which is the difference in volume between $65\mu \mathrm{L}$ and $1.3\mu \mathrm{L}$.

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hCG ER Dose Response Curve

| Conc.
mIU/mL | Rate
mA/min |
| --- | --- |
| 0 | 0.0042 |
| 20 | 0.0812 |
| 80 | 0.3074 |
| 200 | 0.7286 |
| 1000 | 2.9533 |
| 2000 | 5.0696 |

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# Imprecision

Within run and total imprecision were evaluated by testing two commercial controls and three serum pools for twenty days on two systems using two lots of hCG reagent. For both systems, twenty days of data, one run a day, were collected with one reagent lot and 17 days of data were collected with the other reagent lot. The mean rate for each reagent lot was used to generate calibration curves, and concentrations for each lot were calculated from their respective calibration curves. The within-run variance was calculated daily. The among day variance was equal to the variance of the daily means minus the within-run variance divided by the number of reps per day. Total variance was equal to the among day variance plus the within-run variance. Standard deviations were the square roots of the variances.

## Imprecision Results

| Product | Number of Replicates | Mean mIU/mL | Within Run S.D. mIU/mL | Within Run C.V. | Total S.D. mIU/mL | Total C.V. |
| --- | --- | --- | --- | --- | --- | --- |
| Serum Pool 1 | 129 | 978 | 38 | 3.8% | 54 | 5.5% |
| Control 1 | 147 | 9707 | 343 | 3.5% | 585 | 6.2% |
| Control 2 | 130 | 28658 | 1083 | 3.8% | 1926 | 6.8% |
| Serum Pool 2 | 132 | 26746 | 955 | 3.6% | 1262 | 4.8% |
| Serum Pool 3 | 129 | 83756 | 2732 | 3.2% | 5212 | 6.2% |

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# Correlation

104 serum samples were assayed for hCG on the Technicon Immuno 1 with the hCG and hCG ER methods. All samples were assayed undiluted in the hCG ER method. Samples that read between 800 and $1000\mathrm{mIU/mL}$ were assayed undiluted in the standard hCG method. Those that read greater than $1000\mathrm{mIU/mL}$ were manually diluted with Technicon Immuno 1 Sample Diluent B (Product # T03-3574-01) before assaying. Two researchers separately diluted each sample in order to minimize individual biases in dilution.

hCG values ranged from 840 mIU/mL to 88,500 mIU/mL. Results were calculated using linear regression.


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# Interfering Substances/ Specificity

The hCG ER method has the identical reagent and calibrator formulation as the standard hCG method. However the hCG ER method uses $1.3\mu \mathrm{L}$ of sample while the standard method uses $50\mu \mathrm{L}$. With 1/38th the sample volume of the standard method, the hCG ER method has 1/38th the amount of interfering or cross-reacting substances added to the reaction. Assay specificities are expected to be the same as for the standard method.

# Expected Values

The hCG ER method has the same expected values as the standard hCG method. The only difference between the two methods is that the standard hCG method reports hCG values between 0.5 and $1000\mathrm{mIU / mL}$ and the hCG ER method reports hCG values between 800 and $100,000\mathrm{mIU / mL}$.

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# Sample Dilution / Parallelism

For parallelism, three unique serum samples with hCG values close to 100,000 mIU/mL were diluted with Sample Diluent B. The mean observed values were all within 95%-107% of expected.

hCG-ER - Parallelism
Sample A

hCG-ER - Parallelism
Sample B

hCG-ER - Parallelism
Sample C

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# Sample Dilution / Linearity

Three unique pairs of high and low hCG serum samples were co-diluted and assayed. The observed recoveries were all within 99%-107% of expected.




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# Reagent & Calibrator Stability

The same hCG reagent is used for the hCG ER method as for the standard method. No additional stability studies are necessary.

The hCG ER calibrators, Part No. T03-3668-01, are identical in formulation to the Technicon SETpoint Reproductive Calibrators, Part No. T03-3148-01. Shelf life and shipping stability are not necessary.

The hCG ER calibrators are stable for 30 days after reconstitution. Three sets of calibrators were reconstituted and tested individually at Time 0. The three sets were not pooled. Their rates were combined and used for "Week 0" calibration. At weekly intervals only one set of open vial calibrator was tested along with a freshly reconstituted set. A different set was used weekly for five weeks. Serum pool recoveries were calculated from week 0, weekly, and fresh reconstituted calibrations.

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hCG ER Reconstituted Calibrator Stability:
Serum Pool 1

hCG ER Reconstituted Calibrator Stability:
Serum Pool 2

hCG ER Reconstituted Calibrator Stability:
Serum Pool 3

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# Minimal Detectable Concentration

The hCG ER method has a lowest reportable result of 800 mIU/mL. Minimal detectable dose is not determined. The lowest reportable result is used to determine the lower limit of detection rather than minimal detectable dose.