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HomeMILLIPLEX® Multiplex for Luminex® ImmunoassaysQuality is Built into Every MILLIPLEX® Multiplex Kit

Quality is Built into Every MILLIPLEX® Multiplex Kit

For more than 20 years, we have offered the benefits of MILLIPLEX® multiplex immunoassays, containing all the components you need to detect multiple analytes simultaneously. We provide the largest selection of multiplex immunoassays, analytes, and species to meet the needs of investigators across multiple research areas. Rely on the quality we build into each MILLIPLEX® multiplex kit to produce results you trust.

Quality at Every Step

R&D scientist. With over four decades of immunoassay development and manufacturing expertise, MILLIPLEX® kits are developed and quality tested for accurate, reproducible, and biologically relevant analyte measurements.

Expert Research and Development

With over four decades of immunoassay development and manufacturing expertise, all MILLIPLEX® kits are tested for specificity and precision, while Qualified assays are additionally designed for accuracy, reproducibility, and biologically relevant analyte measurement.

Scientific support team. Committed care when and where you need it, with knowledgeable in-house and field-based scientific support for assays and instruments.

Dedicated Support

Committed care when and where you need it, with knowledgeable in-house and field-based scientific support for assays and instruments.

Manufacturing/quality control (QC) scientist. MILLIPLEX® multiplex kits are manufactured in facilities that are ISO 9001:2015 compliant.

Skilled Manufacturing and Quality Control

MILLIPLEX® multiplex kits are manufactured in facilities that are ISO 9001:2015 compliant.

MILLIPLEX® multiplex kit. MILLIPLEX® kit components are produced with quality raw materials and are pre-optimized for easy use and minimal preparation steps.

Optimized Kit Components and Reagents

MILLIPLEX® kit components are produced with quality raw materials and are pre-optimized for easy use and minimal preparation steps.

Screening vs. Qualified MILLIPLEX® Kits

CGI graphic showing a platform with 3 levels. Above the levels is a floating screen showing data with S-curve line graphs and box-and-whisker plots. The top level shows a Millipore kit box with a large number of floating beads. This level represents the MILLIPLEX® screening multiplex panel. A female scientist with a white lab coat is carrying one of the beads down a flight of stairs to the 2nd level, representing how researchers can transition from the screening kits to the MILLIPLEX® qualified multiplex panels. The floating beads are following her down. Next to the stairs is an enlarged xMAP® INTELLIFLEX® instrument and under the stairs is another Millipore kit box. A smaller group of beads can be seen floating on the 2nd level, as well as an enlarged well plate and multi-channel pipette on the sides of the level. Another flight of stairs is seen going to a 3rd bottom level that is slightly hidden. An enlarged single channel pipette and the top of the ultrasensitive SMCxPRO<sup>®</sup> instrument is shown.

Compelling PlexAbility with MILLIPLEX® Screening Assays

  • Profile up to 115 analytes simultaneously
  • Uses same reagents as Qualified assay (no QCs or Serum Matrix)
  • Tested for analyte specificity
  • Ideal for identifying gross differences in experimental groups
  • Sample agnostic
  • Species available: Human

Genuine PlexAbility with MILLIPLEX® Qualified Assays

  • Lot-to-lot reproducibility, rigorous QC testing
  • Includes QCs and Serum Matrix
  • Ideal for long-term or cross-site studies
  • Broadest analyte selection across research areas and species
  • Configurable, premixed, and bulk kit options to fit your exact research needs

MILLIPLEX® Qualified and Screening Assay Development

We define MILLIPLEX® assay specifications during kit development. Verification data is provided in each kit protocol. Every MILLIPLEX® kit includes all the optimized reagents and instructions needed to run the assay and read the assay plate on your Luminex® instrument.

In addition to assay specifications listed in the MILLIPLEX® protocols, we evaluate additional performance criteria during the assay development and verification process, described in Table 1 with example data in Figure 1.

Table 1.MILLIPLEX® assay specifications can be found in the kit protocol or insert or by contacting Technical Support.
Graph A is a S-curve line graph with 2 very similar overlapping curves representing different kit lots with MFI on the y-axis and pg/mL on the x-axis showing that using gold standards as a reference calibrator ensures consistent lot-to-lot standard performance. Graph B is a line graph with 4 different lines each with 3 points representing 4 samples containing IFNα2 with 3 different sample dilutions of 1:2, 1:4, and 1:8. % linearity is on the y-axis and sample dilution is on the x-axis. The graph shows all samples being within the 70-130% range and represents parallelism. Graph C shows 3 S-curve line graphs mostly equally apart from each other with the top line being assay buffer, the middle being matrix 1, and the bottom being matrix 2. MFI is on the y-axis and pg/mL is on the x-axis. Assay buffer shows high recovery, matrix 2 shows low recovery, and matrix 1 shows recovery in range meaning it mimics biological sample conditions. Graph D is a box-and-whisker plot showing a MILLIPLEX® assay compared to 2 different vendor kits in both healthy and disease samples to show method comparison. Y-axis is pg/mL and x-axis are the different kits and sample types for Granzyme B detection. The MILLIPLEX® kit generated concentrations in line with the typical literature ranges while competitors were mostly non-detectable.

Figure 1. MILLIPLEX® assay specifications. A) Gold standards are used as a reference calibrator to ensure consistent standard performance from lot-to-lot. Data generated using Product. No. HCYTA-60K. B) IFNα2 parallelism with calculated results in range of 70-130%. Data generated using Product. No. HIFN-130K. C) Serum Matrix selection ensures accurate measurement by simulating the native sample environment. In this example, the Assay Buffer curve and Matrix 2 curve demonstrate high and low recovery, respectively. Matrix 1 demonstrates recovery in range and mimics biological sample conditions. Data generated using Product. No. HMH3-34K. D) Method comparison is performed to ensure MILLIPLEX® kits are the highest performing assays on the market. In this example, Granzyme B concentrations, generated using Product. No. HCYTB-60K, are in line with literature ranges while the other vendor kit samples were mostly non-detectable. 

Lot-to-Lot Reproducibility In Qualified Assays

Reproducibility across multiple component lots is important to ensure reliable results over time.

See how one of our panels demonstrated superior lot-to-lot consistency of cytokine measurement.

Standard Curves are Checked for Each Analyte Across Kit Lots

Each new lot of MILLIPLEX® standards and quality controls (QCs) are compared to the previous lot and a reference lot, our internal gold standard, to ensure lot-to-lot reproducibility and consistency (Figure 2).

Lot-to-lot comparison of standard curves from MILLIPLEX® multiplex kits. Gold Standard lot compared to three subsequent lots (1K-3K) of the analyte Human IL-6 for Cat. No. HSTCMAG-28SK.

Figure 2. Gold Standard lot compared to three subsequent lots (1K-3K) of the analyte Human IL-6 for Product No. HSTCMAG-28SK.

Additionally, each lot is tested after lyophilization to confirm that there has been no loss due to the lyophilization process. Full standard curve characteristics and relative potency of analytes are maintained within the specifications of the reference lot (Figure 3).

Graph showing point-to-point analysis trend chart for TNFα standard curve MFI values showing consistent values across 85 assays and 4 standard lots.

TNFα Standard Curve Trend Chart

Figure 3. Trend chart standard curve point-to-point analysis shows consistent MFI values for TNFα across 85 assays and 4 standard lots. This data was generated using Product No. HCYTA-60K. Data for one standard curve is shown.

Since MILLIPLEX® panels stand the test of time, new standard lots are periodically assigned to be the fresh reference lot against which subsequent lots are compared. All data is compiled in a single database and trend charts are maintained. Other suppliers compare new standard lots to previous lots, without a reference lot, which makes it difficult to compare data from multiple lots since standard curve point values may vary with each new lot and assay drift may occur.

The Importance of Standard Curve Potency

Analyte standard curves from all lots should overlap as closely as possible. Maintaining standard curve potency acceptance criteria is important because it helps to avoid common problems which negatively impact the calculation of analyte concentration in samples (Figure 4). Problematic standard and component lots are never released for sale.

Graph showing an example of problems that may arise when producing new standard lots, such as curve hooking/prozone effect, curve shift, limited linear range, and curves intersecting, highlighting the importance of standard curve potency.

Figure 4. Example of problems that may arise when producing new standard lots, highlighting the importance of standard curve potency.

Quality Controls to Verify Assay Performance

We include two QCs and QC range sheets to qualify and verify assay performance. Assay results for these QCs are to be checked against the provided QC range sheet values to ensure proper pipetting and assay setup have been accomplished. QC values are based on a minimum of six assays run by at least three different operators. The use of high and low QC values serves as an additional checkpoint in case there was user error associated with hydrating or diluting standards.

Additionally, QCs are important for:

  • Translational studies that require more validation, ensuring that the data are reproducible across kit lots
  • Comparing data across multiple sites
  • Comparing assay results from multiple technicians

As with other MILLIPLEX® components, trend charts are maintained for QC ranges across multiple lots. As an added measure, it is recommended that customers include experiment-relevant sample QCs in each assay (Figure 5).

: 96-well plate layout shows the placement of standards, QCs, and the recommended experiment-relevant sample QCs.

Figure 5. 96-well plate layout shows the placement of standards, QCs, and the recommended experiment-relevant sample QCs.

Other MILLIPLEX® Components

Other components that help enhance the quality of MILLIPLEX® kits include:

Serum Matrix

Because blood is a complex matrix that contains proteins that may interfere with the accurate measurement of the desired analyte, using an optimized serum matrix in the standard curve when measuring analytes secreted in serum or plasma:

  • Significantly improves the accuracy of measurement
  • More accurately simulates the conditions of the native analyte present in serum or plasma
  • Mimics the environment of native analytes in serum or plasma

Bead Diluent

A percentage of the healthy population samples contain heterophilic antibodies that can non-specifically bind to the capture and detection antibodies simultaneously, generating a false positive signal. MILLIPLEX® bead diluents contain a cocktail of proprietary reagents that significantly reduce this false signal without reducing the analyte measurement.

Detection Antibody Cocktail

MILLIPLEX® detection antibody cocktails are designed to yield consistent analyte profiles within the panel, lot-to-lot, regardless of plex size (Figure 6).

Graph showing 2 lines for a singleplex and multiplex assay from the same MILLIPLEX® panel with MFI on the y-axis and ng/mL on the x-axis. It represents consistent analyte profiles.

Consistent Analyte Profile of Properdin

Figure 6. Consistent analyte profiles are seen when comparing multiplex and singleplex assays from the same MILLIPLEX® panel, (Product No. HCMPEX1-19K) as in this example of the analyte Properdin.

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