This in vitro diagnostic procedure is intended to quantitatively measure human chorionic gonadotropin (hCG) in human serum on the Technicon Immuno 1º 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 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℃. 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 nm and 450 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 SET point Reproductive Calibrators, Part No. T03-3148-01, which are used for the standard hCG method. There are no changes in either reagent or callbrator formulation. The hCG ER method differs from the standard hCG method in three ways:

1. The hCG ER calibrators, are reconstituted in 1.0 mL of Reproductive Calibrator Diluent while the Reproductive Calibrators are reconstituted in 2.0mL of Reproductive Calibrator Diluent. This doubles the concentrations of hCG in the calibrator from 0 -1000 mIU/mL to 0 - 2000 mIU/mL.
1. 65uL of calibrator are reacted with reagent, while 1.3uL of any other sample, serum or control, are reacted. 65uL of the highest calibrator, 2000 mIU/mL, corresponds to 130 total mIU of hCG. A corresponding serum sample with 130 total mIU of hCG, at 1.3uL volume, would have 100,000 mIU/mL concentration. Thus a serum sample at 100,000 mIU/mL reacts the same total quantity of hCG, and produces the same amount of p-nitrophenoxide, as does calibrator at 2,000 mIU/mL.
1. The concentrations of all samples other than calibrator are automatically multiplied by a factor of 50, which is the difference in volume between 65uL and 1.3uL.

AI/ML Overview

Here's an analysis of the provided text, focusing on acceptance criteria and supporting study details:

Acceptance Criteria and Device Performance for hCG Extended Range (hCG ER) Method

1. Table of Acceptance Criteria and Reported Device Performance

Performance Metric	Acceptance Criteria (Implied)	Reported Device Performance (hCG ER Method)	Notes
Reference Range	Quantitative measurement of hCG in human serum in an extended range from 800 mIU/mL to 100,000 mIU/mL.	800 - 100,000 mIU/mL	The primary intended use and key differentiation from the standard hCG method.
Precision (Within-run %CV)	Acceptable precision for quantitative diagnostic assay (implied to be similar to standard hCG method's 2.1-2.4% for lower concentrations).	3.2% - 3.8%	While slightly higher than the lower range of the standard method, these values are for significantly higher hCG concentrations, where larger absolute variations may be acceptable. The clinical significance of these CVs at higher concentrations would be evaluated by experts.
Precision (Total %CV)	Acceptable overall precision for quantitative diagnostic assay (implied to be similar to standard hCG method's 3.7-4.0% for lower concentrations).	4.8% - 6.8%	Similar to within-run precision, the values are slightly higher, but for a much higher concentration range.
Correlation with Standard Method (r)	High correlation (implied: >0.95 or similar to demonstrate equivalent performance with the extended range)	0.998 (for 104 sera, range 840-88,500 mIU/mL)	This is an excellent correlation, indicating strong agreement between the hCG ER method and the standard hCG method.
Linearity (Recovery)	Recoveries consistently falling within a defined range (e.g., 90-110% of expected)	99% - 107% of expected	Excellent linearity, demonstrating accurate measurements across the extended range.
Parallelism (Recovery after Dilution)	Mean observed values within a defined range of expected (e.g., 90-110% of expected)	95% - 107% of expected	Good parallelism, indicating that dilution of high samples yields proportional and accurate results. This is critical for assays with an extended range requiring dilution.
Interfering Substances/Specificity	Expected to be the same as the standard hCG method.	Expected to be the same as the standard hCG method due to reduced sample volume.	The claim is based on the reduced sample volume, which theoretically reduces the amount of interfering substances. No new experimental data for specificity is presented for the hCG ER method itself, but relies on the established performance of the standard method.
Expected Values	Same as the standard hCG method.	Same as the standard hCG method.	Confirms that the extended range doesn't alter the clinical interpretation of hCG levels, only the measurable range.
Reagent and Calibrator Stability	Reagent stability for the existing hCG reagent is established. Calibrators stable for 30 days after reconstitution.	Reagent: No additional studies needed. Calibrators: Serum pool recoveries within a target range (implied by "Fresh Recon, Week 0 Calibration, and Weekly Calibration, as well as a -2SD Range line" showing agreement).	Based on the identical formulation to existing products, indicating prior establishment of stability. The calibrator stability study explicitly confirms the 30-day stability of the reconstituted calibrators.
Minimal Detectable Concentration	Lowest reportable result is 800 mIU/mL.	800 mIU/mL (lowest reportable result)	This is a statement of the method's lower limit, not an acceptance criterion to be met by a specific test, but rather a characteristic of the assay design. Minimal detectable dose was not determined, with emphasis on the lowest reportable result for the defined extended range.

2. Sample Sizes and Data Provenance

Test Set for Precision:
- Within-run & Total Precision: n = 130 over 20 days for Controls 1, 2 and Serum Pools 1, 2, 3.
- Data Provenance: Not explicitly stated (e.g., retrospective/prospective, country of origin). It seems to be prospective internal testing by Bayer Corporation as data is "on file at Bayer Corporation".
Test Set for Correlation:
- n = 104 serum samples.
- Data Provenance: Not explicitly stated (e.g., retrospective/prospective, country of origin). Appears to be internal testing.
Test Set for Parallelism:
- 3 unique serum samples.
- Data Provenance: Not explicitly stated.
Test Set for Linearity:
- 3 unique pairs of high and low hCG serum samples.
- Data Provenance: Not explicitly stated.
Test Set for Calibrator Stability:
- 3 sets of calibrators (reconstituted weekly over 5 weeks).
- Data Provenance: Not explicitly stated.

3. Number of Experts and Qualifications for Ground Truth

This submission describes an in vitro diagnostic assay for quantitative measurement of a biomarker (hCG). The ground truth for such assays is typically established by the reference method or established analytical techniques, not by human experts in the sense of image interpretation or clinical diagnosis.
In the correlation study, the "ground truth" (or comparative method) was the standard Technicon Immuno 1 hCG method. This method itself would have been validated against a traceable reference method or gold standard.
For precision, linearity, and parallelism, the "ground truth" is derived from expected values based on the known concentrations of controls/pools or theoretical dilutions, rather than expert consensus on individual samples.

4. Adjudication Method for the Test Set

For an in vitro diagnostic assay evaluating analytical performance like precision, linearity, and correlation, adjudication by multiple human readers (like in imaging studies) is not applicable.
The correlation study involved two researchers separately diluting samples "to minimize individual biases in dilution," which is a form of quality control to reduce measurement error, but not an adjudication of a judgment call.

5. Multi-Reader Multi-Case (MRMC) Comparative Effectiveness Study

No MRMC comparative effectiveness study was done. This type of study is more relevant for diagnostic aids where human interpretation is involved (e.g., radiology AI). This submission focuses on the analytical performance of an automated in vitro diagnostic device.

6. Standalone Performance Study

Yes, this entire submission details the standalone performance of the hCG ER method. It describes the algorithm's (the Immuno 1 system's) ability to quantitatively measure hCG without human interpretation in the loop. The results (precision, correlation, linearity) are direct measurements of the device's output.

7. Type of Ground Truth Used

Reference Method Comparison: For the correlation study, the ground truth was the standard Technicon Immuno 1 hCG method.
Known Concentrations/Expected Values: For precision, linearity, parallelism, and calibrator stability, the ground truth was based on known concentrations in control materials, serum pools, or mathematically derived expected values from dilutions.

8. Sample Size for the Training Set

The document primarily describes the validation or performance verification of the hCG ER method, which is an adaptation of an existing method. It does not explicitly mention a "training set" in the context of machine learning.
The "training" of the system would broadly refer to the calibration of the instrument using a set of calibrators. The document mentions the concentrations of the calibrators: 0, 20, 80, 200, 1000, 2000 mIU/mL (for the hCG ER calibrators). The dose-response curve (page 3) shows these points.

9. How Ground Truth for the Training Set Was Established

The "ground truth" for the calibrators (the "training set" in a broader sense for an IVD) would be established by traceability to a recognized reference standard for hCG, often an International Reference Preparation (IRP) or World Health Organization (WHO) standard. The calibrators are prepared at precisely known concentrations, which are verified against these reference materials. The document states that the hCG ER Calibrators are "identical in formulation" to the Technicon SET point Reproductive Calibrators, suggesting their ground truth is rooted in the same established traceability as the standard method.

Summary

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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

Part Number	TechniconImmuno 1ºhCG ERT01-2966-51	TechniconImmuno 1ºhCGT01-2966-51
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 = Immuno 1 hCG ER X = Immuno 1 hCG n = 104 sera r = 0.998

. .

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Assay Description

1. The hCG ER calibrators, are reconstituted in 1.0 mL of Reproductive Calibrator Diluent while the Reproductive Calibrators are reconstituted in 2.0mL of Reproductive Calibrator Diluent. This doubles the concentrations of hCG in the calibrator from 0 -1000 mIU/mL to 0 - 2000 mIU/mL.
1. 65uL of calibrator are reacted with reagent, while 1.3uL of any other sample, serum or control, are reacted. 65uL of the highest calibrator, 2000 mIU/mL, corresponds to 130 total mIU of hCG. A corresponding serum sample with 130 total mIU of hCG, at 1.3uL volume, would have 100,000 mIU/mL concentration. Thus a serum sample at 100,000 mIU/mL reacts the same total quantity of hCG, and produces the same amount of p-nitrophenoxide, as does calibrator at 2,000 mIU/mL.
1. The concentrations of all samples other than calibrator are automatically multiplied by a factor of 50, which is the difference in volume between 65uL and 1.3uL.

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Image /page/3/Figure/0 description: The image is a plot titled "hCG ER Dose Response Curve". The x-axis is labeled "hCG Concentration (mIU/mL)" and ranges from 0 to 2000. The y-axis is labeled "Reaction Rate (mA/min)" and ranges from 0.00 to 6.00. The plot shows a linear relationship between hCG concentration and reaction rate.

Conc.	Rate
mlU/mL	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.

Product	NumberofReplicates	MeanmlU/mL	Within RunS.D.mlU/mL	Within RunC.V.	TotalS.D.mlU/mL	TotalC.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%

Imprecision Results

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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 mIU/mL were assayed undiluted in the standard hCG method. Those that read greater than 1000 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.

Image /page/5/Figure/3 description: This image is a method comparison of hCG vs. hCG ER on Technicon Immuno 1. The scatter plot shows the correlation between the two methods, with hCG on the x-axis and hCG ER on the y-axis, both measured in mIU/mL. A regression line is plotted through the data points, and the equation of the line is given as hCG = 1.11(hCG ER) + 323, with a correlation coefficient of r = 0.998, based on 104 serum samples.

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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 1.3uL of sample while the standard method uses 50uL. 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 mIU/mL and the hCG ER method reports hCG values between 800 and 100,000 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.

Image /page/7/Figure/2 description: The image contains three graphs, each titled "hCG-ER - Parallelism" with different sample labels: A, B, and C. Each graph plots "Recovery (mIU/mL)" on the y-axis against "Percent Serum" on the x-axis, showing observed data points connected by a dotted line representing the expected values. The graphs include percentage values at each data point, such as 105.0%, 95.8%, 100.1%, and 101.2% for Sample A, indicating the recovery rate at different serum percentages.

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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.

Image /page/8/Figure/2 description: The image contains three line graphs, each titled "HCG-ER LINEARITY" with a different number (#1, #2, and #3). Each graph plots "Recovery (mIU/mL)" on the y-axis against "Linearity Pool" on the x-axis, ranging from -2 to 2. Each graph shows an "Expected" line with "Observed" data points marked with percentages indicating the recovery rate at different linearity pool values, such as 102% at -1, 99% at 0, 101% at 1, and 107% at 2.

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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 SET point 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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Image /page/10/Figure/0 description: The image contains three line plots that show the stability of hCG ER Reconstituted Calibrator for Serum Pools 1, 2, and 3. Each plot shows the recovery in mIU/mL over a period of 5 weeks. The plots include data points for Fresh Recon, Week 0 Calibration, and Weekly Calibration, as well as a -2SD Range line.

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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.

Regulation Number and Section

§ 862.1155 Human chorionic gonadotropin (HCG) test system.

(a)
Human chorionic gonadotropin (HCG) test system intended for the early detection of pregnancy —(1)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.(2)
Classification. Class II.(b)
Human chorionic gonadotropin (HCG) test system intended for any uses other than early detection of pregnancy —(1)Identification. 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.(2)
Classification. Class III.(3)
Date PMA or notice of completion of a PDP is required. As of the enactment date of the amendments, May 28, 1976, an approval under section 515 of the act is required before the device described in paragraph (b)(1) may be commercially distributed. See § 862.3.