Garantirana detekcija in kvantifikacija z uporabo laboratorijsko preverjenih testov Zanesljivi rezultati z integriranimi kontrolami Pripravljena programska orodja za preprosto interpretacijo rezultatov Microbial DNA qPCR Assays (330025) 1X 100 µl tube Microbial DNA qPCR Assay, 1X 1.35 ml tube Microbial qPCR Mastermix Izbor testov: Microbial DNA qPCR Assay/Assay Izbor testovza odpornost na antibiotike: Antibiotic Resistance Gene Resource and Assay List Microbial DNA qPCR Assay Kit (330033) 1X 20 µl tube of Microbial DNA qPCR Assay, 1X tube Positive PCR Control (20 µl), 1X tube Microbial DNA Positive Control (50 µl), 1X tube Microbial DNA-Free Water (1.35 ml), 1X tube Microbial qPCR Mastermix (1.35 ml) Microbial DNA qPCR Arrays (330261) Pred pripravljeni hidrolizirani testi so naneseni na različnih PCR ploščah 96 ( pakiranje 2/12/24), 384 (pakiranje 1/4/16), 100 mestni okrogli nosilec, primernih za različne qPCR naprave. PCR miksi lahko vsebujejo ROX, Fluorescin ali so brez dodatka. Arreji vsebujejo vse reagente za pripravo, tudi vodo brez vsebnosti mikrobne DNA. Testiranje Hrane: Perutnina Testiranje Hrane: Morska hrana Testiranje Hrane: Zelenjava Testiranje Hrane: Mlečni izdelki Testiranje Hrane: Meso Vaginalna flora Bakterijska Vaginoza Okužbe sečil Črevesne okužbe Presnovne motnje Okužbe dihal Ustne bolezni Sepsa Odpornost na antibiotike Analiza vode Biološka obramba Slika: Primer plošče Microbial DNA qPCR Arrays. Na plošči testiramo en vzorec z 90 Microbial testi. Vsaka jamica vsebuje posamezen test. Vključene so tudi kontrole vzorca, pan-bakterijska, pan-glive in pozitivna PCR kontrola Custom Microbial DNA qPCR Arrays (330161) Po želji izbranih 90 testov v želenem vrstnem redu pripravijo na plošče. Hidrolizirani testi so naneseni na različnih ploščah (96(pakiranje 12/24/+12...), 384(pakiranje 6/24/+6...),100 mestni okrogli nosilec ) primernih za različne qPCR naprave. PCR miksi lahko vsebujejo ROX, Fluorescin ali so brez dodatka. Arreji vsebujejo vse reagente za pripravo, tudi vodo brez vsebnosti mikrobne DNA. Minimalno naročilo je 12 (96 mestnih plošč) ali 6 (384 mestnih plošč). Microbial DNA qPCR Multi-Assay Kits (330043) Kompletni kit za mikrobno profiliranje s kontrolami in reagenti Vsebuje ploščo z experimentalno preverjenimi Microbial DNA qPCR aseji. Uporablja se 5' hidrolizirano sondo za občutljivo in specifično detekcijo Microbial DNA qPCR aseji, Microbial DNA Positive Control, Positive PCR Control, Microbial DNA-Free Water, and Microbial qPCR Mastermix. Bartonella henselae Pathogenicity Bordetella pertussis Pathogenicity Brucella spp Pathogenicity Campylobacter jejuni Pathogenicity Chlamydia trachomatis Pathogenicity Clostridium difficile Pathogenicity Clostridium perfringens Pathogenicity C. diphtheriae Pathogenicity Enterococcus faecalis Pathogenicity Enterohaemorrhagic E. coli Pathogenicity Haemophilus influenzae Pathogenicity Helicobacter pylori Pathogenicity Legionella pneumophila Pathogenicity Listeria monocytogenes Pathogenicity Mycobacterium spp Pathogenicity Neisseria meningitidis Pathogenicity Pseudomonas aeruginosa Pathogenicity Salmonella enterica Pathogenicity Shigella dysenteriae Pathogenicity Streptococcus agalactiae Pathogenicity Streptococcus pneumoniae Pathogenicity Streptococcus pyogenes Pathogenicity Vibrio cholerae Pathogenicity Yersinia enterocolitica Pathogenicity Yersinia spp Pathogenicity MRSA: Methicillin-resistant S. aureus Laboratory Rodent Testing SHV Antibiotic Resistance Genes Sestava Microbial DNA qPCR Multi-Assay Kita, MRSA: Methicillin-resistant S. aureus Microbial DNA-Free Water: 338132; Microbial DNA Positive Control: 338134; Supplemental Microbial qPCR Mastermix (Fluorescein): 330540; Supplemental Microbial qPCR Mastermix (ROX): 330530; Tehnične informacije Limit of detection versus lower limit of quantification. This chart demonstrates the difference between the limit of detection (LOD) and the lower limit of quantification (LLOQ). The LOD is defined as the lowest concentration at which 95% of the positive samples are detected, whereas the LLOQ is the lowest concentration that falls within the linear range of a standard curve. LOD depends upon the precision of the assay, and requires at least 40 replicates for determination of a positive sample. For the Microbial DNA qPCR Assays, LLOQ is sufficient to determine assay sensitivity. Linearity and sensitivity of Microbial DNA qPCR Arrays. Linearity and sensitivity for each Microbial DNA qPCR Array was determined using synthetic templates over a 6 log serial dilution ranging from 1 copy to 1 million copies. The following are representative results for all the qPCR assays. [A] shows the realtime amplification curves of the KPC antibiotic resistance gene qPCR assay. In [B], a standard curve was prepared that shows that the primer efficiency equals 103% (calculated from slope = –3.3236) and the correlation coefficient is 0.9983, indicating optimum performance for the KPC qPCR assay. All Microbial DNA qPCR Assays have primer efficiencies between 80–120% and correlation coefficients (R)>0.995. The lower limit of quantification The lower limit of quantification (LLOQ) (LLOQ) for antibiotic resistance gene for all Microbial DNA qPCR detection Microbial DNA qPCR Assays reveals high sensitivity. Assays reveals high sensitivity. This chart shows the distribution of LLOQThis for chart shows the distribution of all Microbial DNA qPCR Assays. 93% of LLOQ all for Microbial DNA qPCR Microbial DNA qPCR Assays have a LLOQ Assays of for antibiotic resistance gene <100 gene copies. detection. 95% of all antibiotic resistance gene assays have a LLOQ of <100 gene copies. The lower limit of quantification (LLOQ) for microbial identification Microbial DNA qPCR Assays reveals high sensitivity. This chart shows the distribution of LLOQ for microbial identification Microbial DNA qPCR Assays. 92% of all microbial identification assays have a LLOQ of <100 gene copies. The lower limit of quantification (LLOQ) for virulence factor gene detection Microbial DNA qPCR Assays reveals high sensitivity. This chart shows the distribution of LLOQ for Microbial DNA qPCR Assays for virulence factor gene detection. 97% of all virulence factor gene assays have a LLOQ of <100 gene copies Microbial DNA qPCR Assays display high sensitivity even in complex metagenomic samples. To ensure that Microbial DNA qPCR Assays performed comparably in a complex sample, where there may be up to a thousand different microbial species, each assay was tested using stool, tooth plaque, and sputum samples. For each sample, synthetic template targets were spiked in and the CT was compared to synthetic template alone. PCR was performed using several sample types, which included pooled synthetic template targets alone, stool, stool plus pooled synthetic template targets, plaque, plaque plus pooled synthetic template targets, sputum, and sputum plus pooled synthetic template targets. If the CT<35 in stool, plaque, or sputum samples alone, then ΔCT was calculated (i.e., CTstool + pooled synthetic template targets – CTpooled synthetic template targets). This calculation was performed for all the assays. For each assay, the ΔCT<3, indicating that a complex metagenomic background does not affect the performance of each Microbial DNA qPCR Assay. Vaginal samples positive for Gardnerella vaginalis also show changes in commensal and bacterial vaginosis-related microbes compared to healthy samples. To compare any differences in the vaginal microbiome between healthy women and women with bacterial vaginosis, each sample that tested positive for Gardnerella vaginalis using the Vaginal Flora Microbial DNA qPCR Array was compared to samples from healthy women (n=3). Fold-change in microbial species abundance was calculated by the ΔΔCT method using human genomic DNA to normalize. The results show that as the relative abundance of Gardnerella vaginalis increases, the abundance of the commensal species Lactobacillus crispatus decreases. Also, an increase in Gardnerella vaginalis was associated with an increase in other bacterial vaginosis-associated microbial species. This suggests that Lactobacillus crispatus protects the vagina from bacterial vaginosis-associated microbial species. Specificity of the Antibiotic Resistance Genes Microbial DNA qPCR Array is verified by pyrosequencing. To verify the specificity of the Antibiotic Resistance Genes Microbial DNA qPCR Array (cat no. BAID-1901Z) results from Klebsiella pneumoniae isolates, pyrosequencing assays were designed to detect for the presence and sequences of SHV-156G, SHV-156D, SHV-238G240E, SHV-238S240K, SHV-238S240E, SHV-238G240K, ermB, mefA, tetA, tetB,CTX-M-1 Group, CTX-M-2 Group, AAC(6′)-lb-cr and aadA1. For each Klebsiella pneumoniae isolate, results from the Antibiotic Resistance Genes Microbial DNA qPCR Array were confirmed by pyrosequencing. Representative pyrograms for [A] SHV-156G, [B] SHV-238/240, [C] KPC and [D] CTX-M-1 group are shown. For SHV variants, the Antibiotic Resistance Gene Microbial DNA qPCR Array was able to reliably distinguish single nucleotide polymorphisms occurring at different sites. The Vaginal Flora Microbial DNA qPCR Array provides accurate profiling for cervical swab samples. The vaginal microbiota is a key component influencing women’s urogenital health. To determine what changes in the vaginal microbiota occurs during bacterial vaginosis, the Vaginal Flora Microbial DNA qPCR Array (cat. no. BAID-1902Z), which detects up to 90 different microbial species, was used to test cervical swabs from healthy individuals and from patients with bacterial vaginosis. Genomic DNA from vaginal samples originating from three patients that tested negative for bacterial vaginosis, three patients that tested positive for Candida, three patients that tested positive for Garderella vaginalis, and one patient that tested positive for Trichomonas vaginalis by BD Affirm™ VPIII Microbial Identification Test were run on the Vaginal Flora Microbial DNA qPCR Array. Genomic DNA from ThinPrep samples were isolated using QIAGEN’s QIAamp MinElute Media Kit and 500 ng genomic DNA from each sample was analyzed. After the PCR run on a Roche LightCycler 480, raw CT values were exported to the Microbial DNA qPCR data analysis software. Positive (+) / negative (blank) / inconclusive (+/-) results for each microbial species were determined using the identification criteria. The results from the Vaginal Flora Microbial DNA qPCR Array were in concordance with the BD Affirm VPIII Microbial Identification Test. The Microbial DNA qPCR Array screens gut microbiota for the presence of antibiotic resistance genes. The human gut microbiota is known to act as a reservoir for antibiotic resistance genes, where they can be transferred horizontally to potential pathogenic bacteria. To detect the presence of antibiotic resistance genes from gut microbiota, stool samples from five healthy adults were collected and genomic DNA was isolated using QIAGEN’s QIAamp DNA Stool Mini Kit. 500 ng genomic DNA from each stool sample was analyzed for presence of antibiotic resistance genes using the Antibiotic Resistance Genes Microbial DNA qPCR Array (cat. no. BAID-1901Z). The Antibiotic Resistance Genes Microbial DNA qPCR Array contains assays for 83 antibiotic resistance genes, assays to identify methicillin-resistant Staphylococcus aureus, and control assays. ErmB, mefA, and tetA were found in all or most of the stool samples tested, showing that they may be highly prevalent in the gut. These antibiotic resistance genes have been reported to be isolated from bacterial strains originating from food, suggesting a possible source of origin. This highlights the importance of increased monitoring of antibiotic resistance reservoirs to identify potential sources of antibiotic-resistant bacteria.
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