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H.6.Investigation: scope of issue and problem statement: customer (b)(6) reported that product 345785 (cd14 pe, clone mop9, ce) lot 9302577 is having performance issue, losing staining efficiency after short time.Customer indicates they used product to analyze thp1 cell line which overexpress cd14 after stimulation.Customer compared flow cytometry results of product 345785 lot 9302577 against another cd14 apc reagent which show brighter staining intensity.Manufacturing defect trend: product 345785 (cd14 pe, clone mop9, ce) lot 9302577 was assembled in bdb cayey (plant 1157) using material subassembly 91-0277 (cd14 pe) batch 9225513 manufactured in plant 1157.Material 91-0277 batch 9225513 was used to manufacture the following materials and corresponding lots: 345785 lot 9302577 (133 ea), 340683 lots 9266537 (75 ea) and 0037462 (65 ea), 347497 lot 9284034 (52 ea).Subassembly 91-0277 batch 9225513 was manufactured according to requirements and met manufacturing and qa acceptance criteria for release.No discrepancies (oos or qn) were identified in manufacturing/testing batch history record (bhr) of subassembly 91-0277 batch 9225513 during evaluated period of 14-jun-21 to 14-jun-20.Complaint trend: only one (1) complaint pr# 3070179-gc for product 345785 (cd14 pe, clone mop9, ce) lot 9302577 or subassembly 91-0277 (cd14 pe) batch 9225513 related to performance issue, losing staining efficiency after short time, have been reported in trackwise during evaluated period of 14-jun-21 to 14-jun-20.It is noted there is no additional complaints reported against product 345785 lot 9302577 from a total of 133 ea sold to market or any other products manufactured (refer to manufacturing defect trend section) using subassembly 91-0277 batch 9225513: 340683 lots 9266537 (75 ea) and 0037462 (65 ea), and 347497 lot 9284034 (52 ea).Batch history record (bhr) review: product 345785 (cd14 pe, clone mop9, ce) lot 9302577 was assembled in plant 1157 using material subassembly 91-0277 (cd14 pe) batch 9225513 manufactured in plant 1157, which met established acceptance criteria for product release thru flow cytometry testing of human peripheral blood, gated on monocytes, as demonstrated by evaluated batch history record (bhr).Manufacturing was performed according to requirements and met specifications without any discrepancy or nonconformance.No discrepancy or deviation was found on the evaluated bhr of subassembly 91-0277 batch 9225513.Product 345785 lot 9302577 expires on 31-aug-21.Retain sample evaluation / testing: retain sample testing not deemed necessary based on evaluation of available bhr, recent test history data and no additional claims for product 345785 (cd14 pe, clone mop9, ce) lot 9302577 or other related reagents/lots manufactured using same subassembly 91-0277 (cd14 pe) batch 9225513 as described on complaints trend section.Bhr review for subassembly 91-0277 batch 9225513 (tested in 2019 for release) found no discrepancy on functional performance results, while further evaluation of qc test history for 91-0277 (cd14 pe) reveal batch 9225513 was later used as reference lot for routine qc test of newer 91-0277 batch 9346687 (tested in 2020), in which flow cytometry results met percent positive and lmf (log mean fluorescence) bd specifications without any discrepancy.Returned sample evaluation: samples were not requested to be returned, since qa retains are kept for subassembly 91-0277 batch 9225513 if needed for further evaluation.Investigation result / analysis: customer reported that product 345785 (cd14 pe, clone mop9, ce) lot 9302577 is having performance issue, losing staining efficiency after short time.Customers provide a picture of flow cytometry data (which was evaluated, as described below) from stained thp1 cell line which overexpress cd14 after in vitro stimulation.As per technical data sheet (tds) or product insert (pi) document 23-5055-04, product 340579 (cd14 pe, clone mop9, ce) is for in vitro diagnostic (ivd).Applications: expression of cd14 in the characterization of hematologic neoplasia.Clone: cd14 antibody, clone mop9, is derived from the hybridization of mouse sp2/0 myeloma cells with spleen cells from balb/c mice immunized with peripheral blood monocytes from a patient with rheumatoid arthritis.Composition: cd14 antibody is composed of mouse igg2b heavy chains and kappa light chains.Each reagent is supplied in pbs containing gelatin and 0.1% sodium azide.Pe reagent volume per test = 20 l.Concentration of reagent = 100 g/ 2.0 ml = 0.05 mg/ml or 0.05 g/l.Recommended reagent volume = 20 l per test, or 20 l (0.05 g/l) = 1.00 g per test.Storage and handling: antibody reagent is stable until the expiration date shown on label when stored at 2-8 °c.Do not use after expiration date.Do not freeze the reagent or expose it to direct light during storage or incubations with cells.Keep outside of the reagent vial dry.Do not use the reagent if any change in appearance is observed.Precipitation indicates instability or deterioration.Specimens: reagent can be used for immunophenotyping by flow cytometry a variety of specimen types, including peripheral blood, bone marrow aspirates or biopsies, and other body fluids or tissues.Each type of specimen can have different storage conditions and limitations that should be considered prior to collection and analysis.Samples with large number of nonviable cells can give erroneous results due to selective loss of populations and to increased nonspecific binding of antibodies to nonviable cells.A cut-off value of at least 80% viable cells has been suggested.Procedure: flow cytometry figure 1 displays representative data (different cd14 antibody-conjugated fluorochromes) performed on whole blood and gated on monocytes.Performance characteristics: specificity the cd14 antigen is present on the majority of normal peripheral blood monocytes.Cd14 has weak reactivity with peripheral blood granulocytes.Cd14 recognizes a human monocyte/macrophage antigen with a molecular weight of 55 kda.Sensitivity is defined as resolution of the cd14+ population from the cd14- population.Each concentration of reagent was tested on whole blood.The bottled antibody concentration for each reagent provided optimum sensitivity in resolving cd14+ cells from the negative.Evaluation of customer provided flow cytometry data results: customer provided flow cytometry results of product 345785 (cd14 pe, clone mop9, ce) lot 9302577 used to stain thp1 cell line and not used to stain human specimens for in vitro diagnosis.Data show product 345785 (cd14 pe, clone mop9, ce) lot 9302577 result was compared against another cd14 apc format reagent, which show relatively brighter staining intensity on their respective fluorescence detection channels.It is known different fluorochrome-conjugated reagents, such as cd14 pe and cd14 apc, may have inherent differences in fluorescence intensity detected due to the nature of the fluorochrome and its relative brightness, assuming product has been properly stored and protected from light, as indicated on (b)(4).Prolonged exposure of fluorochrome-conjugated reagents to out-of-recommended storage conditions and viability or type of stained specimen samples (peripheral blood cells, bone marrow aspirates or biopsies or other tissues) may affect or cause erroneous results.Typically, lot-to-lot variability checks should be assessed using same reagent clone and same fluorochrome-conjugate format.Customer flow cytometry data using product 345785 (cd14 pe, clone mop9, ce) lot 9302577 show expected positive staining on thp1 (activated) target cells versus inactivated thp1 control, however, its fluorescence peak intensity (brightness) is relatively different from that observed with the cd14 apc reagent used.Comparing relative brightness among different fluorochrome format (i.E.Pe vs apc) antibody-conjugated reagents, to assess lot-to-lot variability performance, is not suitable for the reasons explained earlier.Cd14 is a known positive marker on human peripheral blood monocytes, as described and shown on staining profile in (b)(4).Routine qc testing for cd14 clone mop9 subassembly 91-0277 release is performed by flow cytometry, using normal human peripheral blood gated on monocytes population and compared against a reference lot reagent of the same fluorochrome format, per bd specifications.Bd does not have performance characteristics established or historical data for product 345785 (cd14 pe, clone mop9, ce) using thp1 cell line, which requires in vitro activation for cd14 antigen expression.Based on investigation findings for product 345785 (cd14 pe, clone mop9, ce) lot 9302577 or subassembly 91-0277 batch 9225513, in which no discrepancies were found on evaluated bhr or data, no trends showing additional claims reported, and evidence of satisfactory results for 91-0277 (cd14 pe) batch 9225513 when later used as qc reference lot, this claim is not confirmed, and no further actions are deemed necessary at this time.Risk analysis: risk analysis applicable for product 345785 (cd14 pe, clone mop9, ce) is available under risk analysis document fmeacd14mop9, revision 05.Evaluation was based on¿ conjugated antibody ¿ final product¿, which might trigger a problem to customer.Hazard(s) identified? x yes no reviewed item: 1.To detect a specific cell surface marker 2.Not perform to the spec claim.Potential effects of failures: 3.Wrong result ¿ no matching profile, customer inconveniences, additional testing, cost and time.Potential cause of failure: 14.Incorrect volume used @ customer site 18.Improper use of material by the user severity: 7 occurrence: 3 detection: 2 rpn: 42 = acceptable risk current controls: 1.Customer education and 2.Labeling ¿ detailed procedures new hazard: none mitigation(s) enough x yes no root cause analysis: root cause cannot be determined.Customer reported that product 345785 (cd14 pe, clone mop9, ce) lot 9302577, manufactured using subassembly 91-0277 (cd14 pe) batch 9225513, is having performance issue of lower relative fluorescence intensity when compared to another apc-conjugated cd14 reagent.Evidence demonstrates manufacturing process was performed according to requirements; product met specifications and no discrepancy was found.Investigation found subassembly 91-0277 batch 9225513 was later successfully used as qc reference lot for routine qc testing a more recent batch of cd14 pe (91-0277 batch 9346687), obtaining satisfactory results per bd specifications.It is noted customer claim was based on comparing relative brightness among two different fluorochrome format (cd14 pe vs cd14 apc) antibody-conjugated reagents, to assess lot-to-lot variability performance, which is not suitable approach as described on investigation results section.Based on investigation, this claim is not confirmed, and no further actions are deemed necessary at this time.Conclusion: product 345785 (cd14 pe, clone mop9, ce) lot 9302577 manufactured from subassembly 91-0277 (cd14 pe) batch 9225513, was manufactured per requirements and complies with bd specifications.Based on investigation performed, it is determined claim is not confirmed, and no further actions are necessary at this time.
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