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The LIAISON PLEX® Gram-Positive Blood Culture Assay (BCP), performed using the automated, sample-to-result LIAISON PLEX® System, is a qualitative multiplexed in vitro diagnostic test for the simultaneous detection and identification of selected gram-positive pathogens and/or selected genetic determinants associated with antimicrobial resistance in positive blood culture bottles. BCP is performed directly on blood culture media using blood culture bottles identified as positive by a continuous monitoring blood culture system and which contain gram-positive bacteria as determined by Gram stain. The BCP Assay detects and identifies the following: Gram Positive Resistance Markers: - mecA/mecC - vanA - vanB Genera and Species: - Bacillus spp. - Enterococcus faecalis - Enterococcus faecium - Listeria spp. - Staphylococcus spp. - Staphylococcus aureus - Staphylococcus epidermidis - Staphylococcus lugdunensis - Streptococcus spp. - Streptococcus agalactiae - Streptococcus anginosus group - Streptococcus pneumoniae - Streptococcus pyogenes Negative results for antimicrobial resistance genes do not indicate bacterial susceptibility as there are multiple mechanisms that can contribute to resistance. The LIAISON PLEX® BCP Assay contains targets for the detection of genetic determinants associated with resistance to methicillin (mecA/C) and vancomycin (vanA and vanB) to aid in the identification of potentially antimicrobial-resistant organisms in positive blood culture samples. In mixed growth, the LIAISON PLEX BCP Assay does not specifically attribute vanA/vanB-mediated vancomycin resistance to either E. faecalis or E. faecium, or mecA/mecC-mediated methicillin resistance to either Staphylococcus spp., S. aureus, S. epidermidis or S. lugdunensis. The antimicrobial resistance gene or marker detected may or may not be associated with the agent responsible for disease. Negative results for these select antimicrobial resistance gene and marker assays do not indicate susceptibility, as multiple mechanisms of methicillin and vancomycin resistance exist. The LIAISON PLEX® BCP Assay is indicated for use in conjunction with other clinical and laboratory findings to aid in the diagnosis of bacterial bloodstream infections (BSI). The LIAISON PLEX® BCP Assay is not intended to monitor these infections. Sub-culturing of positive blood cultures is necessary to recover organisms for antimicrobial susceptibility testing (AST), for identification of organisms not detected by the LIAISON PLEX BCP Assay, to detect mixed infections that may not be detected by the LIAISON PLEX BCP Assay, for association of antimicrobial resistance genes to a specific organism, or for epidemiological typing.

The LIAISON PLEX® Gram-Positive Blood Culture Assay (BCP Assay) is an automated test for the detection and identification of nucleic acid from gram-positive bacteria in a positive blood culture media sample. The BCP Assay is performed directly on blood culture media using blood culture bottles identified as positive by a continuous monitoring blood culture system, and which contain gram-positive bacteria, as determined by a Gram stain. The LIAISON PLEX® System is a fully automated, bench-top "sample-to-answer" device that performs sample preparation and microarray-based hybridization for the detection of target-specific nucleic acids. The test reagents are supplied as a single, disposable test cartridge. Target amplification is not performed as part of the BCP Assay workflow, as it is a non-amplified, direct detection test performed on the LIAISON PLEX® System.

The LIAISON PLEX® Gram‐Negative Blood Culture (BCN) Assay, performed using the automated, sample‐to‐result LIAISON PLEX® System, is a qualitative multiplexed in vitro diagnostic test for the simultaneous detection and identification of selected gram‐negative pathogens and/or selected genetic determinants associated with antimicrobial resistance in positive blood culture bottles. LIAISON PLEX® BCN Assay is performed directly on blood culture media using blood culture bottles identified as positive by a continuous monitoring blood culture system which contain gram‐negative bacteria as determined by Gram stain. The LIAISON PLEX® BCN Assay detects and identifies the following: Resistance Markers: - CTX‐M (blaCTX‐M) - IMP (blaIMP) - KPC (blaKPC) - NDM (blaNDM) - OXA (blaOXA) - VIM (blaVIM) - MCR - SME (blaSME) Gram Negative Genera and Species: - Enterobacteriaceae / Morganellaceae - Acinetobacter baumannii - Acinetobacter spp. - Citrobacter spp. - Enterobacter spp. (1) - Escherichia coli (2) - Haemophilus influenzae - Klebsiella oxytoca - Klebsiella pneumoniae - Klebsiella variicola - Morganella morganii - Neisseria meningitidis - Proteus spp. - Pseudomonas aeruginosa - Pseudomonas spp. - Salmonella spp. - Serratia marcescens - Stenotrophomonas maltophilia (1) Due to reclassification, Klebsiella aerogenes will be reported Enterobacter spp. (2) LIAISON PLEX® BCN Assay will not distinguish between Escherichia coli and Shigella spp. (S. dysenteriae, S. boydii, S. flexneri and S. sonnei) LIAISON PLEX® BCN Assay contains targets for the detection of genetic determinants associated with resistance to carbapenems (blaCTX‐M, blaIMP, blaKPC, blaNDM, blaOXA48‐like, blaVIM, blaSME) to aid in the identification of potentially antimicrobial‐resistant organisms in positive blood culture samples. In addition, the panel includes an assay for the detection of the mobilized genetic determinant MCR, an emerging marker of public health importance. The antimicrobial resistance gene or marker detected may or may not be associated with the agent responsible for disease. Negative results for these select antimicrobial resistance gene and marker assays do not indicate susceptibility, as multiple mechanisms of resistance to ß‐lactams and colistin exist. LIAISON PLEX® BCN Assay is indicated for use in conjunction with other clinical and laboratory findings to aid in the diagnosis of bacterial bloodstream infections (BSI). LIAISON PLEX® BCN Assay is not intended to monitor treatment of these infections. Sub‐culturing of positive blood cultures is necessary to recover organisms for antimicrobial susceptibility testing (AST), for identification of organisms not detected by LIAISON PLEX® BCN Assay, to detect mixed infections that may not be detected by LIAISON PLEX® BCN Assay, for association of antimicrobial resistance marker genes to a specific organism, or for epidemiological typing.

The LIAISON PLEX® Gram‐Negative Blood Culture (BCN) Assay is an automated test for the detection and identification of nucleic acid from gram‐negative bacteria in a positive blood culture media sample. The BCN Assay is performed directly on blood culture media using blood culture bottles identified as positive by a continuous monitoring blood culture system, and which contain gram‐negative bacteria, as determined by a Gram stain. The LIAISON PLEX® System is a fully automated, bench‐top "sample‐to‐answer" device that performs sample preparation, polymerase chain reaction (PCR) and microarray‐based hybridization for the detection of target‐specific nucleic acids. The test reagents are supplied as a single, disposable test cartridge. PCR is not performed on the LIAISON PLEX® BCN Assay, as it is a non‐amplified, direct detection test performed on the LIAISON PLEX® System.

The BIOFIRE Blood Culture Identification 2 (BCID2) Panel is a multiplexed nucleic acid test intended for use with BIOFIRE FILMARRAY 2.0 or BIOFIRE FILMARRAY TORCH Systems for the simultaneous qualitative detection and identification of multiple bacterial and yeast nucleic acids and select genetic determinants associated with antimicrobial resistance. The BIOFIRE BCID2 Panel test is performed directly on blood culture samples identified as positive by a continuous monitoring blood culture system. Results are intended to be interpreted in conjunction with Gram stain results. The following organism types and subtypes are identified using the BIOFIRE BCID2 Panel: Gram Positive Bacteria Enterococcus faecalis Staphylococcus spp. Streptococcus spp. Enterococcus faecium Staphylococcus aureus Streptococcus agalactiae (Group B) Listeria monocytogenes Staphylococcus epidermidis Streptococcus pneumoniae Staphylococcus lugdunensis Streptococcus pyogenes (Group A) Gram Negative Bacteria Acinetobacter calcoaceticus-baumannii complex Enterobacterales Bacteroides fragilis Enterobacter cloacae complex Haemophilus influenzae Escherichia coli Neisseria meningitidis (encapsulated) Klebsiella aerogenes Pseudomonas aeruginosa Klebsiella oxytoca Stenotrophomonas maltophilia Klebsiella pneumoniae group Proteus spp. Salmonella spp. Serratia marcescens Yeast Candida albicans Candida krusei Cryptococcus neoformans/gattii Candida auris Candida parapsilosis Candida glabrata Candida tropicalis The BIOFIRE BCID2 Panel contains assays for the detection of genetic determinants associated with resistance to methicillin (mecA/C and mecA/C in conjunction with MREJ, vancomycin (vanA and vanB), ß-lactams including penicillins, cephalosporins, monobactams, and carbapenems (blaCTX-M, blaIMP, blaKPC, blaNDM, blaOXA48-like, blaVIM) to aid in the identification of potentially antimicrobial-resistant organisms in positive blood culture samples. In addition, the panel includes an assay for the mobilized genetic determinant mor-1, an emerging marker of public health importance. The antimicrobial resistance gene or may not be associated with the agent responsible for disease. Negative results for these select antimicrobial resistance gene and marker assays do not indicate susceptibility, as multiple mechanisms of resistance to methicillin, vancomycin, b-lactams, and colistin exist. Antimicrobial Resistance Genes CTX-M KPC mecA/C NDM vanA/B IMP mcr-1 mecA/C and MREJ (MRSA) OXA-48-like VIM The BIOFIRE BCID2 Panel is indicated as an aid in the diagnosis of bloodstream infection and results should be used in conjunction with other clinical and laboratory findings. Positive results do not rule out co-infection with organisms not included in the BIOFIRE BCID2 Panel. The BIOFIRE BCID2 Panel is not intended to monitor treatment for bloodstream infection. Subculturing of positive blood cultures is necessary to recover organisms for susceptibility testing and epidemiological typing, to identify organisms in the blood culture that are not detected by the BIOFIRE BCID2 Panel, and for determination of species detected but not identified within complexes, groups, or genera by the BIOFIRE BCID2 Panel assays.

The BIOFIRE Blood Culture Identification 2 (BCID2) Panel is designed to simultaneously identify 43 bacteria and yeast responsible for bloodstream infections, as well as select genetic determinants of antimicrobial resistance (see Table 1), in a timeframe (about an hour) that allows the test results to be used in determining appropriate patient treatment and management. The BIOFIRE BCID2 Panel is performed directly on positive blood culture samples. The BIOFIRE BCID2 Panel is compatible with BioFire's PCR-based in vitro diagnostic BIOFIRE FILMARRAY 2.0 and FILMARRAY TORCH systems for infectious disease testing. A specific software module (i.e., BIOFIRE BCID2 Panel pouch module) is used to perform BIOFIRE BCID2 Panel testing on these systems. A test is initiated by loading Hydration Solution into one port of the BIOFIRE pouch and positive blood culture specimen mixed with the provided Sample Buffer into the other port of the BIOFIRE BCID2 Panel pouch and placing it in a BIOFIRE System. The pouch contains all the reagents required for specimen testing and analysis in a freeze-dried format; the addition of Hydration Solution and Sample/Buffer Mix rehydrates the reagents. After the pouch is prepared, the BIOFIRE Software guides the user though the steps of placing the the instrument, scanning the pouch barcode, entering the sample identification, and initiating the run. The BIOFIRE System contains a coordinated system of inflatable bladders and seal points, which act on the pouch to control the movement of liquid between the pouch blisters. When a bladder is inflated over a reagent blister, it forces liquid from the blister into connecting channels. Alternatively, when a seal is placed over a connecting channel it acts as a valve to open or close a channel. In addition, electronically-controlled pneumatic pistons are positioned over multiple plungers in order to deliver the rehydrated reagents into the blisters at the appropriate times. Two Peltier devices control heating and cooling of the pouch to drive the PCR reactions and the melt curve analysis. Nucleic acid extraction occurs within the BIOFIRE pouch using mechanical and chemical lysis followed by purification using standard magnetic bead technology. After extracting and purifying nucleic acids from the unprocessed sample, the BIOFIRE system performs a nested multiplex PCR that is executed in two stages. During the first stage, the BIOFIRE System performs a single, large volume, highly multiplexed reverse transcription PCR (rt-PCR) reaction. The products from first stage PCR are then diluted with a fresh, primer-free master mix and a fluorescent double stranded DNA binding dye (LC Green® Plus, BioFire Diagnostics). The solution is then distributed to each well of the array. Array wells contain sets of primers designed specifically to amplify sequences internal to the PCR products generated during the first stage PCR reaction. The 2nd stage PCR, or nested PCR, is performed in single plex fashion in each well of the array. At the end of the 2nd stage PCR, the array is interrogated by melt curve analysis for the detection of signature amplicons denoting the presence of specific targets. A digital camera placed in front of the 2nd stage PCR captures fluorescent images of the PCR reactions and software interprets the data. The BIOFIRE Software automatically interprets the results of each DNA melt curve analysis and combines the data with the results of the internal pouch controls to provide a test result for each organism on the panel.

The LIAISON PLEX Yeast Blood Culture (BCY) Assay is a qualitative nucleic acid multiplex in vitro diagnostic test intended for use on the LIAISON PLEX System for simultaneous detection and identification of multiple potentially pathogenic fungal organisms in positive blood culture. The LIAISON PLEX BCY Assay is performed directly on blood culture samples identified as positive by a continuous monitoring blood culture system and which contain fungal organisms as determined by Gram Stain. The LIAISON PLEX BCY Assay detects and identifies the following fungal organisms: Candida albicans Candida auris Candida dubliniensis Candida famata Candida glabrata Candida guilliermondii Candida kefyr Candida krusei Candida lipolytica Candida lusitaniae Candida parapsilosis Candida tropicalis Candida haemulonii / duobushaemulonii Cryptococcus neoformans / gattii The detection and identification of specific fungal nucleic acids from individuals exhibiting signs and/or symptoms of bloodstream infection aids in the diagnosis of bloodstream infection when used in conjunction with other clinical information. The results from LIAISON PLEX BCY Assay are intended to be interpreted in conjunction with Gram stain results and should not be used as the sole basis for diagnosis, treatment management decisions. Negative results in the setting of a suspected bloodstream infection with pathogens that are not detected by this test. Positive results do not rule out co-infection with other organisms; the organism(s) detected by LIAISON PLEX BCY Assay may not be the definite cause of disease. Additional laboratory testing (e.g. sub-culturing of positive blood cultures for identification of organisms not detected by LIAISON PLEX BCY Assay, susceptibility testing and differentiation of mixed growth) and clinical presentation must be taken into consideration in the final diagnosis of bloodstream infection.

The LIAISON PLEX "Yeast Blood Culture Assay (BCY Assay) is performed directly on blood culture media using blood culture bottles identified as positive by a continuous monitoring blood culture system, and which contain a fungal organism, as determined by a Gram stain. The system consists of an instrument, a single-use disposable test cartridge, and a transfer pipette. The user loads the sample into the sample port of the LIAISON PLEX Yeast Blood Culture Assay Cartridge. Next, the user sets up the sample order on the LIAISON PLEX System by first entering the sample information or scanning the barcode ID located on the sample tube, then scanning the barcode ID located on the test cartridge. Last, the user inserts the test cartridge into the processing module to initiate the test. The LIAISON PLEX System identifies the assay being run and automatically initiates the proper testing protocol to process the sample, analyze the data, and generate test results. The LIAISON PLEX System automates the BCY Assay sample analysis through the following steps: a) Sample Preparation: Nucleic acid extraction via mechanical and chemical cell lysis and magnetic bead-based nucleic acid isolation; b) Amplification: Multiplex PCR based amplification of the extracted nucleic acid to generate target specific amplicons; c) Hybridization: Amplified DNA hybridizes to specific capture DNA arrayed on a glass slide in a microarray format and the bound target DNA, in turn, hybridizes with mediator and gold-nanoparticle probes; d) Signal Analysis: Gold nanoparticle probes bound specifically to target-containing spots in the microarray are silver-enhanced, and light scatter from the spots is measured and further analyzed to determine the presence (Detected) or absence (Not Detected) of a target.

The GenMark ePlex® Blood Culture Identification Gram-Negative (BCID-GN) Panel is a qualitative nucleic acid multiplex in vitro diagnostic test intended for use on GenMark's ePlex Instrument for simultaneous qualitative detection and identification of multiple potentially pathogenic gram-negative bacterial organisms and select determinants associated with antimicrobial resistance in positive blood culture. In addition, the ePlex BCID-GN Panel is capable of detecting several gram-positive bacteria (Pan Gram-Positive assay) and several Candida species (Pan Candida assay). The ePlex BCID-GN Panel is performed directly on blood culture samples identified as positive by a continuous monitoring blood culture system and which contain gram-negative organism. The following bacterial organisms and genes associated with antibiotic resistance are identified using the ePlex BCID-GN Panel: Acinetobacter baumannii, Bacteroides fragilis, Citrobacter sakazakii, Enterobacter cloacae complex, Enterobacter (non-cloacae complex), Escherichia coli, Fusobacterium necrophorum, Fusobacterium nucleatum, Haemophilus influenzae, Klebsiella oxytoca, Klebsiella pneumoniae group, Morganii, Neisseria meningitidis, Proteus, Proteus mirabilis, Pseudomonas aeruginosa, Salmonella, Serratia marcescens, Stenotrophomonas maltophilia, CTX-M (blaCTX-M), IMP (blaMP) , KPC (blaKPC) , NDM (blaNDM), OXA (blaOXA) (OXA-23 and OXA-48 groups only), and VIM (blaVIM). The ePlex BCID-GN Panel contains assays for the detection of genetic determinants associated with resistance to antimicrobial agents including CTX-M(blaCTX-M), which is associated with resistance to extended spectrum betalactamase (ESBL)-mediated resistance to penicillins, cephalosporins, and monobactams, as well as OXA (blaOXA) (OXA-23 and OXA-48 groups only), KPC (blaKPC), and metallo-beta-lactamases IMP (blaIMP), and NDM (blaNDM), which is associated with carbapenemase-mediated resistance. The antimicrobial resistance gene detected may or may not be associated with the agent responsible for disease. Negative results for these select antimicrobial resistance assays do not indicate susceptibility, as there are multiple mechanisms of resistance in gramnegative bacteria. The ePlex BCID-GN Panel also contains targets designed to detect a broad range of organisms with a potentially misleading Gram stain result or organisms that may be missed by Gram staining altogether, for example in the case of coinfections. These include a broad Pan Gram-Positive assay (which is designed to detect Bacillus cereus group, Bacillus subtilis group, Enterococcus, Staphylococus, and Streptococcus), as well as a Pan Candida assay, which is designed to detect four Candida species: Candida albicans, Candida krusei, and Candida parapsilosis. The detection and identification of specific bacterial and fungal nucleic acids from individuals exhibiting signs and/or symptoms of bloodstream infection aids in the diagnosis of bloodstream infection when used in conjunction with other clinical information. The results from the ePlex BCID-GN Panel are intended to be interpreted in conjunction with Gram stain results and should not be used as the sole basis for diagnosis, treatment, or other patient management decisions. Negative results in the setting of a suspected bloodstream infection with pathogens that are not detected by this test. Positive results do not rule out co-infection with other organisms; the organism(s) detected by the ePlex BCID-GN Panel may not be the definite cause of disease. Additional laboratory testing (e.g. sub-culturing of positive blood cultures for identification of organisms not detected by ePlex BCID-GN Panel and for susceptibility testing, differentiation of mixed growth, and association of antimicrobial resistance marker genes to a specific organism) and clinical presentation must be taken into consideration in the final diagnosis of bloodstream infection.

The ePlex Blood Culture Identification Gram-Negative (BCID-GN) Panel is based on the principles of competitive nucleic acid hybridization using a sandwich assay format, wherein a single-stranded target binds concurrently to a sequence-specific solution-phase signal probe and a solid-phase electrode-bound capture probe. The test employs nucleic acid extraction, target amplification via polymerase chain reaction (PCR) or reverse transcription PCR (RT-PCR) and hybridization of target DNA. In the process, the double-stranded PCR amplicons are digested with exonuclease to generate single-stranded DNA suitable for hybridization. Nucleic acid extraction from biological samples occurs within the cartridge via cell lysis, nucleic acid capture onto magnetic beads, and release for amplification. The nucleic acid extraction is processed through microfluidic liquid handling. Once the nucleic acid targets are captured and inhibitors are washed away, the magnetic particles are delivered to the electrowetting environment on the printed circuit board (PCB) and the targets are eluted from the particles and amplified. During hybridization, the single-stranded target DNA binds to a complementary, single-stranded capture probe immobilized on the working gold electrode surface. Single-stranded signal probes (labeled with electrochemically active ferrocenes) bind to specific target sequence / region adjacent to the capture probe. Simultaneous hybridization of target to signal probes and capture probe is detected by alternating current voltammetry (ACV). Each working electrode on the array contains specific capture probes, and sequential analysis of each electrode allows detection of multiple analyte targets.

The BioFire® Blood Culture Identification 2 (BCID2) Panel is a multiplexed nucleic acid test intended for use with FilmArray® 2.0 or FilmArray® Torch systems for the simultaneous qualitative detection and identification of multiple bacterial and yeast nucleic acids and select genetic determinants associated with antimicrobial resistance. The BioFire BCID2 Panel test is performed directly on blood culture samples identified as positive by a continuous monitoring blood culture system. Results are intended to be interpreted in conjunction with Gram stain results. The following organism types and subtypes are identified using the BioFire BCID2 Panel: Gram Positive Bacteria - Enterococcus faecalis - · Staphylococcus spp. - · Streptococcus spp. - Enterococcus faecium - Staphylococcus aureus - · Streptococcus agalactiae (Group B) - Listeria monocytogenes - Staphylococcus epidermidis - Streptococcus pneumoniae - Staphylococcus lugdunensis - · Streptococcus pyogenes (Group A) Gram Negative Bacteria - Acinetobacter calcoaceticus-baumannii complex - · Enterobacterales - · Bacteroides fragilis - Enterobacter cloacae complex - Haemophilus influenza - · Escherichia coli - · Neisseria meningitidis (encapsulated) - · Klebsiella aerogenes - · Pseudomonas aeruginosa - · Klebsiella oxytoca - · Stenotrophomonas maltophilia - · Klebsiella pneumoniae group - · Proteus spp. - · Salmonella spp. - · Serratia marcescens Yeast - · Candida albicans - Candida krusei - · Cryptococcus neoformans/gattii - Candida auris - · Candida parapsilosis - · Candida tropicalis - Candida glabrata The BioFire BCID2 Panel contains assays for the detection of genetic determinants associated with resistance to methicillin (mecA/C and mecA/C in conjunction with MREJ, vancomycin (vanA and vanB), 0-lactams including penicillins, cephalosporins, monobactams, and carbapenems (blaCTX-M, blaKPC, blaNDM, blaOXA48-like, bla VIM) to aid in the identification of potentially antimicrobial-resistant organisms in positive blood culture samples. In addition, the panel includes an assay for the mobilized genetic determinant mcr-1, an emerging marker of public health importance. The animicrobial resistance gene or may not be associated with the agent responsible for disease. Negative results for these select antimicrobial resistance gene and marker assays do not indicate susceptibility, as multiple mechanisms of resistance to methicillin, vancomycin, B-lactams, and colistin exist. Antimicrobial Resistance Genes - CTX-M - КРС - · mecA/C - NDM - vanA/B - · IMP - mcr-1 - · mecA/C and MREJ (MRSA) - OXA-48-like - VIM The BioFire BCID2 Panel is indicated as an aid in the diagnosis of bloodstream infection and results should be used in conjunction with other clinical and laboratory findings. Positive results do not rule out co-infection with organisms not included in the BioFire BCID2 Panel is not intended to monitor treatment for bloodstream infection. Subculturing of positive blood cultures is necessary to recover organisms for susceptibility testing and epidemiological typing, to identify organisms in the blood culture that are not detected by the BioFire BCID2 Panel, and for determination of species detected but not identified within complexes, groups, or genera by the BioFire BCID2 Panel assays.

The BioFire Blood Culture Identification 2 (BCID2) Panel is designed to simultaneously identify 43 bacteria and yeast responsible for bloodstream infections, as well as select genetic determinants of antimicrobial resistance (see Table 1), in a timeframe(about an hour) that allows the test results to be used in determining appropriate patient treatment and management. The BioFire BCID2 Panel is performed directly on positive blood culture samples. The BioFire BCID2 Panel is compatible with BioFire's PCR-based in vitro diagnostic FilmArray Torch systems for infectious disease testing, A specific software module (i.e., BioFire BCID2 Panel pouch module) is used to perform BioFire BCID2 Panel testing on these systems. A test is initiated by loading Hydration Solution into one port of the FilmArray pouch and positive blood culture specimen mixed with the provided Sample Buffer into the other port of the BioFire BCID2 Panel pouch and placing it in a FilmArray instrument. The pouch contains all of the reagents required for specimen testing and analysis in a freeze-dried format: the addition of Hydration and Sample/Buffer Mix rehydrates the reagents. After the pouch is prepared, the FilmArray Software quides the user through the pouch into the instrument, scanning the pouch barcode, entering the sample identification, and initiating the run. The FilmArray instruments contain coordinated systems of inflatable bladders and seal points, which act on the pouch to control the movement of liquid between the pouch blisters. When a bladder is inflated over a reagent blister, it forces liquid from the blister into connecting channels. Alternatively, when a seal is placed over a connecting channel it acts as a valve to open or close a channel. In addition, electronically-controlled pneumatic pistons are positioned over multiple plungers in order to deliver the rehydrated reagents into the blisters at the appropriate times. Two Peltier devices control heating and cooling of the PCR reactions and the melt curve analysis. Nucleic acid extraction occurs within the FilmArray pouch using mechanical lysis followed by purification using standard magnetic bead technology. After extracting and purifying nucleic acids from the unprocessed sample, the FilmArray performs a nested multiplex PCR that is executed in two stages. During the first stage, the FilmArray performs a single, large volume, highly multiplexed reverse transcription PCR (rt-PCR) reaction. The products from first stage PCR are then diluted and combined with a fresh, primer-free master mix and a fluorescent double-stranded DNA binding dye (LC Green® Plus, BioFire Diagnostics). The solution is then distributed to each well of the array. Array wells contain sets of primers designed specifically to amplify sequences internal to the PCR products generated during the first stage PCR reaction. The 2nd stage PCR, is is performed in singleplex fashion in each well of the conclusion of the 2nd stage PCR, the array is interrogated by melt curve analysis for the detection of signature amplicons denoting the presence of specific targets. A digital camera placed in front of the 2nd stage PCR captures fluorescent images of the PCR reactions and software interprets the data. The FilmArray Software automatically interprets the results of each DNA melt curve analysis and combines the data with the results of the internal pouch controls to provide a test result for each organism on the panel.

The iCubate, Inc. iC-GN Assay™ for use on the iC-System™ is a qualitative, multiplexed, in vitro diagnostic test for the detection and identification of potentially pathogenic gram negative bacteria, which may cause bloodstream infection (BSI). The iC-GN Assay™ is performed directly on positive blood cultures, confirmed by Gram stain to contain gram negative bacilli. Cultures demonstrating mixed Gram stain results should not be tested on the assay. The iC-GN Assay™ is validated for use with select BACTEC™, BacT/ALERT® and VersaTREK® blood culture bottles. The iC-GN Assay™ is indicated for use in conjunction with other clinical and laboratory findings, such as culture, to aid in the diagnosis of bacterial bloodstream infections; however, it is not used to monitor bloodstream infections. The iC-GN Assay™ detects target DNA and identifies the following: Bacterial Genera and Species: Acinetobacter baumannii complex, Enterobacter cloacae complex, Escherichia coli, Klebsiella oxytoca, Klebsiella pneumoniae, Pseudomonas aeruginosa, Proteus species, Serratia marcescens Resistance Markers: KPC (blaKPC)- associated with resistance to carbapenems, NDM (blaNDM)- associated with resistance to carbapenems, CTX-M group 1(blaCTX-M group 1)- associated with resistance to extended spectrum beta-lactams In mixed growth, the iC-GN Assay™ does not specifically attribute detection of KPC, NDM, or CTX-M group 1 to a specific genera or species. Sub-culturing of positive blood cultures is necessary to recover organisms for susceptibility testing, identification of organisms not detected by the iC-GN Assay™, differentiation of mixed growth, association of antimicrobial resistance marker genes to a specific organism, or for epidemiological typing.

The iC-GN Assay™ utilizes polymerase chain reaction (PCR) for the multiplex amplification of specific targets and detects the amplified targets with microarray hybridization. Targets are detected directly from patient positive blood cultures confirmed by Gram stain to contain gram negative bacilli. The iC-GN Assay utilizes proprietary ARM-PCR (Amplicon Rescued Multiplex PCR) technology allowing for multiple targets to be amplified in one reaction. Testing is done in a self-contained, automated, disposable cassette using the iCubate™ processor (iC-Processor™). After the reaction is complete, the cassette is read on the iCubate® reader (iC-Reader™). Results from the iC-Reader™ are interpreted by iC-Report™ software and a final report is displayed on the iMac® computer. To operate, the user opens the iC-Cassette™ cap and pipettes an aliquot of the diluted positive blood culture sample into the sample/PCR well in the bottom well plate of the cassette. Once inoculated, the cassette cap is closed, and all extraction, amplification and detection processes are completed in the cassette, a closed system. Extraction, amplification and detection sequences are defined by an assay script controlled by the iC-Processor™. The processing script is defined within a barcode label positioned on the top of each iC-Cassette™ which communicates with the iC-Processor™. To access and pierce the foilsealed reagent wells located in the bottom well plate of the cassette, the processor manipulates the cassette to move the cassette pipette horizontally and vertically. The script directs the transfer of reagents between the wells in the bottom well plate and finally to the array within the cassette. The iC-Processor™ is capable of processing four (4) iC-Cassettes™ with random access. Once processing is complete, the cassette is manually transferred from the iC-Processor™ to the iC-Reader™ where the microarray within the cassette is read. The iC-Reader™ is capable of reading up to four (4) iC-Cassettes™ at one time. The results are interpreted via the iC-Report™ software and displayed for the user on the iMac®. Raw data and result interpretations are stored within the iMac®; raw data is accessible to iCubate® service personnel only and not to the end user. When finished with a loaded iC-GN Cassette™, it should be disposed as biohazardous waste.

The GenMark ePlex® Blood Culture Identification Gram-Negative (BCID-GN) Panel is a qualitative nucleic acid multiplex in vitro diagnostic test intended for use on GenMark's ePlex Instrument for simultaneous qualitative detection and identification of multiple potentially pathogenic gram-negative bacterial organisms and select determinants associated with antimicrobial resistance in positive blood culture. In addition, the ePlex BCID-GN Panel is capable of detecting several gram-positive bacteria (Pan Gram-Positive assay) and several Candida species (Pan Candida assay). The ePlex BCID-GN Panel is performed directly on blood culture samples identified as positive by a continuous monitoring blood culture system and which contain gram-negative organism. The following bacterial organisms and genes associated with antibiotic resistance are identified using the ePlex BCID-GN Panel: Acinetobacter baumannii, Bacteroides fragilis, Citrobacter, Cronobacter sakazakii. Enterobacter cloacae complex, Enterobacter (non-cloacae complex), Escherichia coli, Fusobacterium necrophorum, Fusobacterium nucleatum, Haemophilus influenzae, Klebsiella oxytoca, Klebsiella pneumoniae group, Morganella morganii, Neisseria meningitidis, Proteus, Proteus mirabilis, Pseudomonas aeruginosa, Salmonella, Serratia, Serratia marcescens, Stenotrophomas maltophilia, СТХ-М (blactх-м), IMP (blамм) , КРС (blakec) , NDM (bland), OXA (blaoxa) (OXA-23 and OXA-48 groups only), and VIM (blaviм). The ePlex BCID-GN Panel contains assays for the detection of genetic determinants associated with resistance to antimicrobial agents including CTX-M(blactx.M), which is associated with resistance to extended spectrum beta-lactamase (ESBL)-mediated resistance to penicillins, cephalosporins and monobactams, as well as OXA (blaoxA) (OXA-23 and OXA-48 groups only), KPC (blakpc), and metallo-beta-lactamases IMP (blavM), VIM (blavM), and NDM (blaNDM), which is associated with carbapenemase-mediated resistance. The antimicrobial resistance gene detected may or may not be associated with the agent responsible for disease. Negative results for these select antimicrobial resistance assays do not indicate susceptibility, as there are multiple mechanisms of resistance in gram-negative bacteria. The ePlex BCID-GN Panel also contains targets designed to detect a broad range of organisms with a potentially misleading Gram stain result or organisms that may be missed by Gram staining altogether, for example in the case of co-infections. These include a broad Pan Gram-Positive assay (which is designed to detect Bacillus cereus group, Bacillus subtilis group, Enterococcus, Staphylococcus, and Streptococcus), as well as a Pan Candida assay, which is designed to detect four Candida species: Candida albicans, Candida glabrata, Candida krusei, and Candida parapsilosis. The detection and identification of specific bacterial and fungal nucleic acids from individuals exhibiting signs and/or symptoms of bloodstream infection aids in the diagnosis of bloodstream infection when used in conjunction with other clinical information. The results from the ePlex BCID-GN Panel are intended to be interpreted in conjunction with Gram stain results and should not be used as the sole basis for diagnosis, treatment, or other patient management decisions. Negative results in the setting of a suspected bloodstream infection may be due to infection with pathogens that are not detected by this test. Positive results do not rule out co-infection with other organisms; the organism(s) detected by the ePlex BCID-GN Panel may not be the definite cause of disease. Additional laboratory testing (e.g. sub-culturing of positive blood cultures for identification of organisms not detected by ePlex BCID-GN Panel and for susceptibility testing, differentiation of mixed growth, and association of antimicrobial resistance marker genes to a specific organism) and clinical presentation must be taken into consideration in the final diagnosis of bloodstream infection.

The ePlex Blood Culture Identification Gram-Negative (BCID-GN) Panel is based on the principles of competitive nucleic acid hybridization using a sandwich assay format, wherein a single-stranded target binds concurrently to a sequence-specific solution-phase signal probe and a solid-phase electrode-bound capture probe. The test employs nucleic acid extraction, target amplification via polymerase chain reaction (PCR) or reverse transcription PCR (RT-PCR) and hybridization of target DNA. In the process, the double-stranded PCR amplicons are digested with exonuclease to generate single-stranded DNA suitable for hybridization. Nucleic acid extraction from biological samples occurs within the cartridge via cell lysis, nucleic acid capture onto magnetic beads, and release for amplification. The nucleic acid extraction is processed through microfluidic liquid handling. Once the nucleic acid targets are captured and inhibitors are washed away, the magnetic particles are delivered to the electrowetting environment on the printed circuit board (PCB) and the targets are eluted from the particles and amplified. During hybridization, the single-stranded target DNA binds to a complementary, single-stranded capture probe immobilized on the working gold electrode surface. Single-stranded signal probes (labeled with electrochemically active ferrocenes) bind to specific target sequence / region adjacent to the capture probe. Simultaneous hybridization of target to signal probes and capture probe is detected by alternating current voltammetry (ACV). Each working electrode on the array contains specific capture probes, and sequential analysis of each electrode allows detection of multiple analyte targets.

The GenMark ePlex Blood Culture Identification Fungal Pathogen (BCID-FP) Panel is a qualitative nucleic acid multiplex in vitro diagnostic test intended for use on GenMark's ePlex Instrument for simultaneous detection and identification of multiple potentially pathogenic fungal organisms in positive blood culture. The ePlex BCID-FP Panel is performed directly on blood culture samples identified as positive by a continuous monitoring blood culture system and which contain fungal organism. The following fungal organisms are identified using the ePlex BCID-FP Panel: Candida albicans, Candida auris, Candida dubliniensis, Candida famata, Candida glabrata, Candida guilliermondii, Candida kefyr, Candida krusei, Candida lusitaniae, Candida parapsilosis, Candida tropicalis, Cryptococcus gattii, Cryptococcus neoformans, Fusarium and Rhodotorula. The detection and identification of specific fungal nucleic acids from individuals exhibiting signs and/or symptoms of bloodstream infection aids in the diagnosis of bloodstream infection when used in conjunction with other clinical information. The results from the ePlex BCID-FP Panel are intended to be interpreted in conjunction with Gram stain results and should not be used as the sole basis for diagnosis, treatment, or other patient management decisions. Negative results in the setting of a suspected bloodstream infection may be due to infection with pathogens that are not detected by this test. Positive results do not rule out co-infection with other organisms; the organism(s) detected by the ePlex BCID-FP Panel may not be the definite cause of disease. Additional laboratory testing (e.g. sub-culturing of positive blood cultures for identification of organisms not detected by ePlex BCID-FP Panel, susceptibility testing and differentiation of mixed growth) and clinical presentation must be taken into consideration in the final diagnosis of bloodstream infection.

The ePlex Blood Culture Identification Fungal Pathogen (BCID-FP) Panel is based on the principles of competitive nucleic acid hybridization using a sandwich assay format, wherein a single-stranded target binds concurrently to a sequence-specific solution-phase signal probe and a solid-phase electrode-bound capture probe. The test employs nucleic acid extraction, target amplification via polymerase chain reaction (PCR) or reverse transcription PCR (RT-PCR) and hybridization of target DNA. In the process, the double-stranded PCR amplicons are digested with exonuclease to generate single-stranded DNA suitable for hybridization. Nucleic acid extraction from biological samples occurs within the cartridge via cell lysis, nucleic acid capture onto magnetic beads, and release for amplification. The nucleic acid extraction is processed through microfluidic liquid handling. Once the nucleic acid targets are captured and inhibitors are washed away, the magnetic particles are delivered to the electrowetting environment on the printed circuit board (PCB) and the targets are eluted from the particles and amplified. During hybridization, the single-stranded target DNA binds to a complementary, single-stranded capture probe immobilized on the working gold electrode surface. Single-stranded signal probes (labeled with electrochemically active ferrocenes) bind to specific target sequence / region adjacent to the capture probe. Simultaneous hybridization of target to signal probes and capture probe is detected by alternating current voltammetry (ACV). Each working electrode on the array contains specific capture probes, and sequential analysis of each electrode allows detection of multiple analyte targets.

The GenMark ePlex Blood Culture Identification Gram-Positive (BCID-GP) Panel is a qualitative nucleic acid multiplex in vitro diagnostic test intended for use on GenMark's ePlex Instrument for simultaneous qualitative detection and identification of multiple potentially pathogenic gram-positive bacterial organisms and select determinants associated with antimicrobial resistance in positive blood culture. In addition, the ePlex BCID-GP Panel is capable of detecting a wide variety of gram-negative bacteria (Pan Gram-Negative assay) and several Candida species (Pan Candida assay). The ePlex BCID-GP Panel is performed directly on blood culture samples identified as positive by a continuous monitoring blood culture system and which contain gram-positive organism. The following bacterial organisms and genes associated with antibiotic resistance are identified using the ePlex BCID-GP Panel: Bacillus cereus group, Bacillus subtilis group, Corynebacterium, Cutibacterium acnes (Propionibacterium acnes), Enterococcus, Enterococcus faecalis, Enterococcus faecium, Lactobacillus, Listeria monocytogenes, Micrococcus, Staphylococcus, Staphylococcus aureus, Staphylococcus epidermidis, Staphylococcus lugdunensis, Streptococcus agalactiae (GBS), Streptococcus anginosus group, Streptococcus pneumoniae, Streptococcus pyogenes (GAS), mecA, mecC, vanA and vanB. The ePlex BCID-GP Panel contains assays for the detection of genetic determinants associated with resistance to methicillin (mecA and mecC) to aid in the identification of potentially antimicrobial resistant organisms in positive blood culture samples. The antimicrobial resistance gene detected may or may not be associated with the agent responsible for disease. The ePlex BCID-GP Panel also contains targets designed to detect a broad range of organisms with a potentially misleading Gram stain result or organisms that may be missed by Gram staining altogether, for example in the case of co-infections. These include a broad Pan Gram-Negative assay as well as a Pan Candida assay, which is designed to detect four of the most prevalent Candida species: Candida albicans, Candida glabrata, Candida krusei and Candida parapsilosis. The detection and identification of specific bacterial and fungal nucleic acids from individuals exhibiting signs and/or symptoms of bloodstream infection aids in the diagnosis of bloodstream infection when used in conjunction with other clinical information. The results from the ePlex BCID-GP Panel are intended to be interpreted in conjunction with Gram stain results and should not be used as the sole basis for diagnosis, treatment, or other patient management decisions. Negative results in the setting of a suspected bloodstream infection may be due to infection with pathogens that are not detected by this test. Positive results do not rule out co-infection with other organisms; the organism(s) detected by the ePlex BCID-GP Panel may not be the definite cause of disease. Additional laboratory testing (e.g. sub-culturing of positive blood cultures for identification of organisms not detected by ePlex BCID-GP Panel and for susceptibility testing. differentiation of mixed growth and association of antimicrobial resistance marker genes to a specific organism) and clinical presentation must be taken into consideration in the final diagnosis of blood stream infection.

The ePlex Blood Culture Identification Gram-Positive (BCID-GP) Panel is based on the principles of competitive nucleic acid hybridization using a sandwich assay format, wherein a single-stranded target binds concurrently to a sequence-specific solution-phase signal probe and a solid-phase electrode-bound capture probe. The test employs nucleic acid extraction, target amplification via polymerase chain reaction (PCR) or reverse transcription PCR (RT-PCR) and hybridization of target DNA. In the process, the double-stranded PCR amplicons are digested with exonuclease to generate single-stranded DNA suitable for hybridization. Nucleic acid extraction from biological samples occurs within the cartridge via cell lysis, nucleic acid capture onto magnetic beads, and release for amplification. The nucleic acid extraction is processed through microfluidic liquid handling. Once the nucleic acid targets are captured and inhibitors are washed away, the magnetic particles are delivered to the electrowetting environment on the printed circuit board (PCB) and the targets are eluted from the particles and amplified. During hybridization, the single-stranded target DNA binds to a complementary, single-stranded capture probe immobilized on the working gold electrode surface. Single-stranded signal probes (labeled with electrochemically active ferrocenes) bind to specific target sequence / region adjacent to the capture probe. Simultaneous hybridization of target to signal probes and capture probe is detected by alternating current voltammetry (ACV). Each working electrode on the array contains specific capture probes, and sequential analysis of each electrode allows detection of multiple analyte targets.