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FilmArray, an automated nested multiplex PCR system for multi-pathogen detection: development and application to respiratory tract infection.

Poritz MA, Blaschke AJ, Byington CL, Meyers L, Nilsson K, Jones DE, Thatcher SA, Robbins T, Lingenfelter B, Amiott E, Herbener A, Daly J, Dobrowolski SF, Teng DH, Ririe KM - PLoS ONE (2011)

Bottom Line: FilmArray has the potential to detect greater than 100 different nucleic acid targets at one time.Validation of the FilmArray technology was achieved through development of a panel of assays capable of identifying 21 common viral and bacterial respiratory pathogens.We demonstrate that automated identification of pathogens from their corresponding target amplicon(s) can be accomplished by analysis of the DNA melting curve of the amplicon.

View Article: PubMed Central - PubMed

Affiliation: Idaho Technology, Inc., Salt Lake City, Utah, United States of America. mark_poritz@idahotech.com

ABSTRACT
The ideal clinical diagnostic system should deliver rapid, sensitive, specific and reproducible results while minimizing the requirements for specialized laboratory facilities and skilled technicians. We describe an integrated diagnostic platform, the "FilmArray", which fully automates the detection and identification of multiple organisms from a single sample in about one hour. An unprocessed biologic/clinical sample is subjected to nucleic acid purification, reverse transcription, a high-order nested multiplex polymerase chain reaction and amplicon melt curve analysis. Biochemical reactions are enclosed in a disposable pouch, minimizing the PCR contamination risk. FilmArray has the potential to detect greater than 100 different nucleic acid targets at one time. These features make the system well-suited for molecular detection of infectious agents. Validation of the FilmArray technology was achieved through development of a panel of assays capable of identifying 21 common viral and bacterial respiratory pathogens. Initial testing of the system using both cultured organisms and clinical nasal aspirates obtained from children demonstrated an analytical and clinical sensitivity and specificity comparable to existing diagnostic platforms. We demonstrate that automated identification of pathogens from their corresponding target amplicon(s) can be accomplished by analysis of the DNA melting curve of the amplicon.

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Related in: MedlinePlus

FilmArray pouch.(A) A FilmArray pouch was injected with mock sample (here colored blue for illustrative purposes) in the left side injection port and hydration solution (colored red) in the right side injection port. (B) The blisters of a FilmArray pouch were filled with different coloring (and the channels between the blisters heat sealed shut). In this pouch the plunger tree was made from plastic dyed blue. The fitment and film are normally at right angles to each other; for clarity the pouch has been flattened. (C) A schematic of the pouch showing a trace of the blisters, channels and array wells (black) and the functional areas of the pouch (red).
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pone-0026047-g001: FilmArray pouch.(A) A FilmArray pouch was injected with mock sample (here colored blue for illustrative purposes) in the left side injection port and hydration solution (colored red) in the right side injection port. (B) The blisters of a FilmArray pouch were filled with different coloring (and the channels between the blisters heat sealed shut). In this pouch the plunger tree was made from plastic dyed blue. The fitment and film are normally at right angles to each other; for clarity the pouch has been flattened. (C) A schematic of the pouch showing a trace of the blisters, channels and array wells (black) and the functional areas of the pouch (red).

Mentions: Each FilmArray pouch is comprised of an injection molded polypropylene reservoir (the “fitment”, 120 mm long, 10 mm wide, 25 mm high, “A” in Figure 1B) heat welded to two sheets of a polyester/polypropylene film containing a copolymer adhesive layer. The sheets of film are welded together using heated plates to form the pattern of channels and “blisters” (“C” through “H” in Figure 1B) comprising the sample processing stations and an area containing a 102-well array for the second stage PCR. The fitment contains 12 reservoirs (6 mm inner diameter on 9 mm spacing) that hold the biochemical reagents. During pouch manufacture three additional reagents are inserted into the appropriate blisters of the pouch and the film is sealed shut under vacuum. Ceramic beads are inserted into the sample lysis blister (“C” in Figure 1). A lyophilized pellet of silica-magnetic beads is inserted into blister “E” (Figure 1). A lyophilized pellet of the oligonucleotides (Idaho Technology, Inc. (ITI)) used in the first stage multiplex PCR is inserted into blister “G”.


FilmArray, an automated nested multiplex PCR system for multi-pathogen detection: development and application to respiratory tract infection.

Poritz MA, Blaschke AJ, Byington CL, Meyers L, Nilsson K, Jones DE, Thatcher SA, Robbins T, Lingenfelter B, Amiott E, Herbener A, Daly J, Dobrowolski SF, Teng DH, Ririe KM - PLoS ONE (2011)

FilmArray pouch.(A) A FilmArray pouch was injected with mock sample (here colored blue for illustrative purposes) in the left side injection port and hydration solution (colored red) in the right side injection port. (B) The blisters of a FilmArray pouch were filled with different coloring (and the channels between the blisters heat sealed shut). In this pouch the plunger tree was made from plastic dyed blue. The fitment and film are normally at right angles to each other; for clarity the pouch has been flattened. (C) A schematic of the pouch showing a trace of the blisters, channels and array wells (black) and the functional areas of the pouch (red).
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3198457&req=5

pone-0026047-g001: FilmArray pouch.(A) A FilmArray pouch was injected with mock sample (here colored blue for illustrative purposes) in the left side injection port and hydration solution (colored red) in the right side injection port. (B) The blisters of a FilmArray pouch were filled with different coloring (and the channels between the blisters heat sealed shut). In this pouch the plunger tree was made from plastic dyed blue. The fitment and film are normally at right angles to each other; for clarity the pouch has been flattened. (C) A schematic of the pouch showing a trace of the blisters, channels and array wells (black) and the functional areas of the pouch (red).
Mentions: Each FilmArray pouch is comprised of an injection molded polypropylene reservoir (the “fitment”, 120 mm long, 10 mm wide, 25 mm high, “A” in Figure 1B) heat welded to two sheets of a polyester/polypropylene film containing a copolymer adhesive layer. The sheets of film are welded together using heated plates to form the pattern of channels and “blisters” (“C” through “H” in Figure 1B) comprising the sample processing stations and an area containing a 102-well array for the second stage PCR. The fitment contains 12 reservoirs (6 mm inner diameter on 9 mm spacing) that hold the biochemical reagents. During pouch manufacture three additional reagents are inserted into the appropriate blisters of the pouch and the film is sealed shut under vacuum. Ceramic beads are inserted into the sample lysis blister (“C” in Figure 1). A lyophilized pellet of silica-magnetic beads is inserted into blister “E” (Figure 1). A lyophilized pellet of the oligonucleotides (Idaho Technology, Inc. (ITI)) used in the first stage multiplex PCR is inserted into blister “G”.

Bottom Line: FilmArray has the potential to detect greater than 100 different nucleic acid targets at one time.Validation of the FilmArray technology was achieved through development of a panel of assays capable of identifying 21 common viral and bacterial respiratory pathogens.We demonstrate that automated identification of pathogens from their corresponding target amplicon(s) can be accomplished by analysis of the DNA melting curve of the amplicon.

View Article: PubMed Central - PubMed

Affiliation: Idaho Technology, Inc., Salt Lake City, Utah, United States of America. mark_poritz@idahotech.com

ABSTRACT
The ideal clinical diagnostic system should deliver rapid, sensitive, specific and reproducible results while minimizing the requirements for specialized laboratory facilities and skilled technicians. We describe an integrated diagnostic platform, the "FilmArray", which fully automates the detection and identification of multiple organisms from a single sample in about one hour. An unprocessed biologic/clinical sample is subjected to nucleic acid purification, reverse transcription, a high-order nested multiplex polymerase chain reaction and amplicon melt curve analysis. Biochemical reactions are enclosed in a disposable pouch, minimizing the PCR contamination risk. FilmArray has the potential to detect greater than 100 different nucleic acid targets at one time. These features make the system well-suited for molecular detection of infectious agents. Validation of the FilmArray technology was achieved through development of a panel of assays capable of identifying 21 common viral and bacterial respiratory pathogens. Initial testing of the system using both cultured organisms and clinical nasal aspirates obtained from children demonstrated an analytical and clinical sensitivity and specificity comparable to existing diagnostic platforms. We demonstrate that automated identification of pathogens from their corresponding target amplicon(s) can be accomplished by analysis of the DNA melting curve of the amplicon.

Show MeSH
Related in: MedlinePlus