(458 days)
Ortho-mune OK-COMBO CD4-FITC/CD8-PE is intended for use in identification and enumeration of CD4+ and CD8+ human T lymphocytes in whole blood by flow cvtometry. The intended use is the same as the intended use of the predicate device. Simultest CD4/CD8 (Leu-3a/2a) commercially distributed by Becton Dickinson Immunocytometry Systems.
Ortho-mune OK-COMBO CD4-FITC/CD8-PE (OKT4A/OKT8) Monoclonal Antibody (Murine) is a blend of the individual purified monoclonal antibodies OKT4A and OKT8 conjugated to the fluorochromes fluorescein isothiocyanate and phycoerythrin respectively.
1. Table of Acceptance Criteria and Reported Device Performance
| Test Type | Acceptance Criteria | Reported Device Performance |
|---|---|---|
| Equivalence | Demonstrate equivalence to the predicate device (Simultest CD4/CD8) in identification and enumeration of CD4+ and CD8+ human T lymphocytes in whole blood. | Regression Analysis:- CD4+: Y = 1.886 + 1.08500 (Ortho-mune) vs. Predicate. Correlation coefficient R = 0.971.- CD8+: Y = 0.112 + 0.84400 (Ortho-mune) vs. Predicate. Correlation coefficient R = 0.955.Mean % Positive Stained Cells (Normal Donors, N=190):- CD4+: Ortho-mune: 47.5% (Range: 11.6-70.8); Predicate: 44.1% (Range: 13.1-61.2)- CD8+: Ortho-mune: 27.8% (Range: 10.9-72.8); Predicate: 29.5% (Range: 12.5-69.1)Mean % Positive Stained Cells (AIDS/ARC Patients, N=83):- CD4+: Ortho-mune: 18.2% (Range: 0.1-60.4); Predicate: 15.8% (Range: 0.2-50.4)- CD8+: Ortho-mune: 56.1% (Range: 12.6-81.9); Predicate: 58.9% (Range: 22.5-88.4)Conclusion: Performance shown to be equivalent to the predicate device. |
| Reproducibility | Acceptable within and between laboratory reproducibility for determining total CD4+ and CD8+ lymphocyte percentages. | Within Laboratory Reproducibility (N=11 donors, 3 sites):- CD4+ Low (mean 11.232%): CVs from 6.701% to 8.268%- CD4+ Normal (mean 49.455%): CVs from 2.443% to 3.118%- CD4+ High (mean 67.528%): CVs from 2.093% to 3.360%- CD8+ Low (mean 7.521%): CVs from 7.186% to 15.735%- CD8+ Normal (mean 32.782%): CVs from 3.102% to 4.244%- CD8+ High (mean 56.234%): CVs from 2.462% to 3.392%Between Laboratory Reproducibility (N=11 donors, 3 sites):- CD4+ Low: CV 5.496%- CD4+ Normal: CV 1.189%- CD4+ High: CV 1.729%- CD8+ Low: CV 5.453%- CD8+ Normal: CV 0.427%- CD8+ High: CV 1.354%Conclusion: Acceptable within and between laboratory reproducibility demonstrated. |
| Linearity | Demonstrate linear performance for both total CD4+ and CD8+ lymphocyte subsets across a range of lymphocyte counts. | Linear performance demonstrated for both total CD4+ and CD8+ lymphocyte subsets across a lymphocyte count range of 20 cells/ul to 18,676 cells/ul. Slopes were indistinguishable from 1 and R values were 1.000. |
2. Sample Sizes Used for the Test Set and Data Provenance
-
Test Set for Equivalence (Comparison with Predicate Device):
- Sample Size: 190 normal donors and 83 AIDS/ARC patients (total 273 specimens).
- Data Provenance: The study was conducted at "three external, geographically distinct sites." The countries of origin are not specified, but the use of "normal donors" and "AIDS/ARC patients" suggests clinical samples. The data is prospective, collected specifically for this study.
-
Test Set for Reproducibility:
- Sample Size: 11 normal donors. Samples from these donors were processed to create low, normal, and high relative percent CD4+ and CD8+ cells. Each sample was stained in replicates of 10.
- Data Provenance: Not explicitly stated, but samples were processed and analyzed at "three independent laboratories." This implies the samples were real human whole blood specimens. The data is prospective.
-
Test Set for Linearity:
- Sample Size: 4 normal donors. Specimens were processed to produce samples with low, normal, and high numbers of lymphocyte subsets. Each sample was stained in triplicate.
- Data Provenance: Not explicitly stated, but implies real human whole blood specimens. The data is prospective.
3. Number of Experts Used to Establish the Ground Truth for the Test Set and Qualifications of Those Experts
Not applicable. This device is an in vitro diagnostic reagent for flow cytometry, which measures cell populations. The "ground truth" for this type of device is the measurement obtained from the predicate device and the internal consistency/characteristics (reproducibility, linearity) of the new device itself. There are no human expert adjudicators for establishing a "ground truth" in the way one would for diagnostic imaging. The comparison is against an established method and assessed by statistical measures.
4. Adjudication Method for the Test Set
Not applicable. As described above, this is a quantitative measurement device, not an imaging or qualitative diagnostic device requiring expert adjudication. The comparison with the predicate device was based on statistical regression analysis and direct comparison of mean percentages and ranges.
5. If a Multi-Reader Multi-Case (MRMC) Comparative Effectiveness Study Was Done, If So, What Was the Effect Size of How Much Human Readers Improve with AI vs Without AI Assistance
Not applicable. This is not an AI/imaging device that involves human readers or interpretation of complex cases. It is an in vitro diagnostic reagent that identifies and enumerates specific cell populations using flow cytometry.
6. If a Standalone (i.e. algorithm only without human-in-the-loop performance) Was Done
Yes, in a sense. The "device" (reagent) is evaluated in a standalone manner by comparing its output (percentage of positive stained cells) directly against that of a predicate device and by assessing its intrinsic performance characteristics (reproducibility, linearity). The analysis is performed by a flow cytometer, and the interpretation of the results from the flow cytometer (i.e., the percentages) is then compared between the new reagent and the predicate reagent. There is no "human-in-the-loop performance" that would alter the quantitative output of the reagent and cytometer system itself.
7. The Type of Ground Truth Used
- For Equivalence Study: The "ground truth" was established by the predicate device's measurements (Simultest CD4/CD8). The study aimed to show that the new device's measurements were statistically equivalent to the predicate device's measurements.
- For Reproducibility and Linearity Studies: The "ground truth" was established based on internal consistency and expected biological behavior. For reproducibility, the ground truth is that repeated measurements of the same sample should yield similar results (low variance). For linearity, the ground truth is that the measured cell counts should scale proportionally with dilution.
8. The Sample Size for the Training Set
Not applicable. This device is a monoclonal antibody reagent used in flow cytometry, not a machine learning or AI model. Therefore, there is no "training set" in the context of algorithm development. Its performance is based on the inherent specificity and binding characteristics of the antibodies.
9. How the Ground Truth for the Training Set Was Established
Not applicable, as there is no training set for this type of device.
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REVISED 510(k) SUMMARY
| SUBMITTER: | Ortho Diagnostic Systems1001 U.S. Hwy 202Raritan, NJ 08869-0606 | CONTACT: | Joanne HarrisTel: (908) 218-8404Fax: (908) 218-8168 |
|---|---|---|---|
| DEVICE NAME: | Ortho-mune™ OK-COMBOCD4-FITC/CD8-PE(OKT™4A/OKT8)Monoclonal Antibody (Murine) | PREDICATE: | Simultest™ CD4/CD8(Leu™-3a/2a) |
| DATE: | February 16, 1996 |
DEVICE DESCRIPTION
Ortho-mune OK-COMBO CD4-FITC/CD8-PE (OKT4A/OKT8) Monoclonal Antibody (Murine) is a blend of the individual purified monoclonal antibodies OKT4A and OKT8 conjugated to the fluorochromes fluorescein isothiocyanate and phycoerythrin respectively.
INTENDED USE
Ortho-mune OK-COMBO CD4-FITC/CD8-PE is intended for use in identification and enumeration of CD4+ and CD8+ human T lymphocytes in whole blood by flow cvtometry. The intended use is the same as the intended use of the predicate device. Simultest CD4/CD8 (Leu-3a/2a) commercially distributed by Becton Dickinson Immunocytometry Systems.
TECHNOLOGICAL CHARACTERISTICS
Both Ortho-mune OK-COMBO CD4-FITC/CD8-PE (OKT4A/OKT8) Monoclonal Antibody (Murine) and Simultest CD4/CD8(Leu-30/2a) utilize monoclonal antibodies specific for human helper/inducer T cells (OKT4A/Leu 3a) and human suppressor/cytotoxic T cells (OKT8/Leu-2a) respectively, conjugated to the same fluorochromes, fluorescein isothiocvanate and phycoerythrin.
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PERFORMANCE CHARACTERISTICS
Performance of the two color reagent Ortho-mune OK-COMBO CD4-FITC/CD8-PE (OKT4A/OKT8) Monoclonal Antibody (Murine) was compared with that of Simultest CD4/CD8 (Leu-3a/2a) at three external, geographically distinct sites. Whole blood specimens from 190 normal donors, and 83 AIDS/ARC patients were stained and analyzed using the ORTHO CYTORONABSOLUTE™ flow cytometer, Ortho Diagnostic Systems Inc.
For each specimen, the percentage of gated cells which showed positive by each marker was calculated. The mean and range of the percent CD4+ and percent CD8+ cells for the normal donor and AIDS/ARC population are shown in Table 1 and Table 2 respectively.
| PERCENT POSITIVE STAINED CELLS IN NORMAL DONORS DETECTEDBY OKT4A/OKT8 AND LEU-3a/2a ASSAYED ON THE CYTORONABSOLUTEN=190 | |||||
|---|---|---|---|---|---|
| Ortho-muneReagent | Mean % | Range % | BDReagent | Mean % | Range % |
| CD4+(OKT4A) | 47.5 | 11.6-70.8 | CD4+(LEU3a) | 44.1 | 13.1-61.2 |
| CD8+(OKT8) | 27.8 | 10.9-72.8 | CD8+(LEU2a) | 29.5 | 12.5-69.1 |
TABLE 1
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| TABLE | 1 |
|---|---|
| ra A | ાં આવેલું એક ગામના લોકોનો મુખ્ય વ્યવસાય ખેતી, ખેતમજૂરી તેમ જ પશુપા |
| PERCENT POSITIVE STAINED CELLS IN AIDS/ARC PATIENTS DETECTEDBY OKT4A/OKT8 AND LEU-3a/2a ASSAYED ON THE CYTORONABSOLUTEN=83 | |||||
|---|---|---|---|---|---|
| Ortho-muneReagent | Mean % | Range % | BDReagent | Mean % | Range % |
| CD4+(OKT4A) | 18.2 | 0.1-60.4 | CD4+(LEU3a) | 15.8 | 0.2-50.4 |
| CD8+(OKT8) | 56.1 | 12.6-81.9 | CD8+(LEU2a) | 58.9 | 22.5-88.4 |
Linear regression analysis of total percent CD4+ cells from the combined normal and AIDS/ARC populations is found in Chart 1. Likewise, linear regression analysis of percent CD8+ cells from the combined normal and AIDS/ARC populations is found in Chart 2.
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CHART 1
OK-COMBO CD4/CD8 vs Simultest Leu-3a/2a
TOTAL CD4+ PERCENT
Image /page/3/Figure/2 description: This image is a scatter plot that compares the OK-COMBO Reagent to the Predicate Reagent. The x-axis represents the Predicate Reagent, and the y-axis represents the OK-COMBO Reagent. The plot shows a strong positive correlation between the two reagents. The equation of the line of best fit is Y = 1.886 + 1.08500, and the correlation coefficient R is 0.971.
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CHART 2
OK-COMBO CD4/CD8 vs Simultest Leu-32/2a
Image /page/4/Figure/2 description: This image is a scatter plot that compares the OK-COMBO Reagent and Predicate Reagent. The x-axis represents the Predicate Reagent, and the y-axis represents the OK-COMBO Reagent. The data points are clustered around a line, which indicates a positive correlation between the two reagents. The equation of the line is Y = 0.112 + 0.84400, and the R-value is 0.955.
TOTAL CD8+ PERCENT
These studies demonstrate that the performance of the two color reagent Ortho-mune OK-COMBO CD4-FITC/CD8-PE (OKT4A/OKT8) Monoclonal Antibody (Murine) is equivalent to Simultest CD4/CD8 (Leu-3a/2a) reagent in identification and enumeration of CD4+ and CD8+ human lymphocytes in whole blood by flow cytometry.
Ortho Diagnostic Systems Inc. Ortho-mune OK-COMBO CD4-FITC/CD8-PE Ref. No. K95-0625 Additional Information Submitted February, 1996 Page 132
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Reproducibility studies were performed at three independent laboratories using samples with low, normal, and high relative vercent CD4+ and CD8+ cells
Specimens from each of eleven normal donors (whole blood, EDT A) were processed using monoclonal antibodies bound to microbeads to produce samples of low, normal, and high relative percent CD4+ and CD8+ cells. The samples were separated into aliquots for each laboratory. Samples were stained in replicates of 10 with Ortho-mone OK-COMBO CD4/CD8 reagent and analyzed using the ORTHO CYTORONABSOLUTE flow cvtometer.
For within laboratory reproducibility, the variance for the replicate results was calculated within site, concentration and donor. The variance was averaged across site, concentration and donor. The square root replicate variance (SD) was divided by the appropriate mean percent positive result (by site and concentration) and multiplied by 100 to obtain the CV. Within laboratory reproducibility results for determination of total percent CD4+ and percent CD8+ cells are presented in Table 3
| WITHIN LABORATORY REPRODUCIBILITY | |||||||
|---|---|---|---|---|---|---|---|
| OK-COMBO CD4/CD8 | |||||||
| N = 11 donors | |||||||
| All SITESMeanPercentPositive | Site A | Site B | Site C | ||||
| OK-COMBOCD4/CD8 | CV | #Reps | CV | #Reps | CV | #Reps | |
| TOTAL CD4+ Low | 11.232 | 8.268 | 98 | 7.358 | 109 | 6.701 | 109 |
| TOTAL CD4+ Normal | 49.455 | 3.118 | 110 | 2.443 | 110 | 2.736 | 109 |
| TOTAL CD4+ High | 67.528 | 3.049 | 110 | 2.093 | 110 | 3.360 | 104 |
| TOTAL CD8+ Low | 7.521 | 15.735 | 110 | 7.186 | 110 | 7.972 | 104 |
| TOTAL CD8+ Normal | 32.782 | 4.244 | 110 | 3.102 | 110 | 3.442 | 109 |
| TOTAL CD8+ High | 56.234 | 3.392 | 98 | 2.462 | 109 | 2.740 | 109 |
TABLE 3
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The between laboratory CV was computed as follows. The mean percent positive for each site within concentration was calculated. The SD was computed on the three site means within concentration and the CV was obtained by dividing the SD by the overall mean within concentration and multiplying by 100. Between laboratory reproductifity results for deternination of total percent CDA+ and percent CD8+ cells are presented in Table 4.
TABLE 4
| BETWEEN LABORATORY REPRODUCIBILITY | |||||
|---|---|---|---|---|---|
| OK-COMBO CD4/CD8 | |||||
| N= 11 donors | |||||
| OK-COMBOCD4/CD8 | SITE AMean Percent Positive(All Donors) | SITE BMean Percent Positive(All Donors) | SITE CMean Percent Positive(All Donors) | ACROSS SITECoefficientofVariation | #Reps |
| TOTAL CD4+ Low | 10.744 | 11.994 | 11.389 | 5.496 | 316 |
| TOTAL CD4+ Normal | 50.103 | 49.392 | 48.937 | 1.189 | 329 |
| TOTAL CD4+ High | 67.934 | 68.473 | 66.235 | 1.729 | 324 |
| TOTAL CD8+ Low | 7.908 | 7.623 | 7.097 | 5.453 | 324 |
| TOTAL CD8+ Normal | 32.636 | 32.913 | 32.741 | 0.427 | 329 |
| TOTAL CD8+ High | 55.979 | 56.787 | 55.271 | 1.354 | 316 |
Ortho-mune OK-COMBO CD4/CD8 immunophenotyping reagent shows acceptable within and between laboratory reproducibility for determination of total CD4+ and CD8+ lymphocyte percentages
Ortho Diagnostic Systems Inc. Ortho-mune OK-COMBO CD4-FTTC/CD8-PE Ref. No. K95-062.5 Additional Information Submitted February, 1996 Page 134
159
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A linearity study was performed using an automated hematology analyzer to determine total lymphocyte count, and the CYTORONABSOLUTE flow cytometer to determine the percent positive CDx cells.
Specimens from four normal donors (whole blood, EDTA) were processed to produce samples with low, normal and high numbers of lymphocyte subsets. Each whole blood specimen was concentrated by harvesting the buffy cost to obtain a white blood cell count between 20,000 and 40,000 cells/ul and then diluting to produce samples of high, normal and low numbers of lymphocyte subsets. A portion of each sample was stained in triplicate using Ortho-muse OK-COMBO CD4/CDS immunophenotyping reagent and analyzed using the CYTORONABSOLUTE flow cytometer. The total lymphocyte count of the concentrated sample for each donor was obtained using an automated hematology analyzer.
Linear regression analyses were performed as follows. The expected (X axis ) values were calculated by multiplying the corresponding serial dilutions by the hematology analyzer derived buffy coat lymphocyte count and by the CYTORONABSOLUTE derived lymphocyte subset percent positive. The observed (Y axis) values were determined as the total lymphocyte count calculated from the hematology derived value of the concentrated sample times the CYTORONABSOLUTE derived lymphocyte subset percent positive at each dilution.
The OK-COMBO CD4/CD8 reagent demonstrated linear performance for both total CD4+ and CD8+ lymphocvte subsets across a lymphocyte count range of 20 celle/ul to 18,676 cells/ul as demonstrated with slopes indistinguishable from 1 and R values of 1.000.
Linear regression analyses of observed versus expected values for total percent CD4+ and percent CD8+ cells for each donor specimen are shown in Chart 3 and Chart 4 respectively. Regression analysis statistics are provided in Table 5.
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Image /page/8/Figure/0 description: The image is a graph titled "OK-COMBO CD4/CD8 CD4+". The x-axis is labeled "Expected CD4+ Counts" and ranges from 0 to 10000. The y-axis is labeled "Observed CD4+ Counts" and ranges from 0 to 10000. The graph shows a linear relationship between the expected and observed CD4+ counts.
CHART 3
DONOR *** 71 8-3 ପ 72 *** 73 合 ★ ★ 74
CHART 4 OK - COMBO CO4CD8 CD8+
Image /page/8/Figure/3 description: This image is a graph with two axes labeled "Expected CD3+ Counts" and "Observed CD3+ Counts". The x-axis, "Expected CD3+ Counts", ranges from 0 to 8000, while the y-axis, "Observed CD3+ Counts", ranges from 0 to 9000. A dashed line runs through the graph, starting at the origin and ending at approximately (6000, 6000). There are several points plotted on the graph, which appear to follow the trend of the dashed line.
士士 ★ 74 DONOR *** 71 8-0 - 72 ++ 73
Ortho Diagnostic Systems Inc.
Ortho-muse OK-COMBO CD4-FITC/CD8-PE Ref. No. K95-0625 Additional Information Submitted February, 1996 Page 136
161
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CONCLUSION
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Performance of the two color reagent Ortho-mune OK-COMBO CD4-FITC/CDS-PE (OKT4A/OKTB) Monoclonal Antibody (Murine) is equivalent to Becton Dickinson (CA.14AOK To) Monocrolar at novement for idemification and desembination of percent CD4+ and CD8+ tymphocytes in whole blood by Bow wyssenses .
Ortho Diagnonnie Sjonamic Itar Cortho-mune ON «CCMMBO CD4-FITC/CD8-PE Ref. You W205-0625 Additional Internantine Submitted February, 1996 Page 137
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్రాల్య
§ 864.5220 Automated differential cell counter.
(a)
Identification. An automated differential cell counter is a device used to identify one or more of the formed elements of the blood. The device may also have the capability to flag, count, or classify immature or abnormal hematopoietic cells of the blood, bone marrow, or other body fluids. These devices may combine an electronic particle counting method, optical method, or a flow cytometric method utilizing monoclonal CD (cluster designation) markers. The device includes accessory CD markers.(b)
Classification. Class II (special controls). The special control for this device is the FDA document entitled “Class II Special Controls Guidance Document: Premarket Notifications for Automated Differential Cell Counters for Immature or Abnormal Blood Cells; Final Guidance for Industry and FDA.”