Analysis‐only detection of <i>Giardia</i> by combining immunomagnetic separation and two‐color flow cytometry

Ferrari, Belinda C.; Veal, Duncan

doi:10.1002/cyto.a.10009

Cited by 21 publications

(20 citation statements)

References 26 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…These include both flow cytometry-based techniques and solid-phase image scanning cytometry techniques (3,4,(6)(7)(8). The latter usually uses a planar multielement photodetector, such as a charge-coupled device (CCD) camera, to make images from adjacent elements across the specimen, followed by scene segmentation and use of pattern recognition algorithms to locate the particular targets (9)(10)(11)(12)(13)(14)(15).…”

mentioning

confidence: 99%

Automated detection of rare‐event pathogens through time‐gated luminescence scanning microscopy

Lu¹,

Jin²,

Leif³

et al. 2011

Cytometry Pt A

View full text Add to dashboard Cite

Many microorganisms have a very low threshold (\10 cells) to trigger infectious diseases, and, in these cases, it is important to determine the absolute cell count in a lowcost and speedy fashion. Fluorescent microscopy is a routine method; however, one fundamental problem has been associated with the existence in the sample of large numbers of nontarget particles, which are naturally autofluorescent, thereby obscuring the visibility of target organisms. This severely affects both direct visual inspection and the automated microscopy based on computer pattern recognition. We report a novel strategy of time-gated luminescent scanning for accurate counting of rare-event cells, which exploits the large difference in luminescence lifetimes between the lanthanide biolabels, [100 ls, and the autofluorescence backgrounds, \0.1 ls, to render background autofluorescence invisible to the detector. Rather than having to resort to sophisticated imaging analysis, the background-free feature allows a single-element photomultiplier to locate rare-event cells, so that requirements for data storage and analysis are minimized to the level of image confirmation only at the final step. We have evaluated this concept in a prototype instrument using a 2D scanning stage and applied it to rare-event Giardia detection labeled by a europium complex. For a slide area of 225 mm 2 , the time-gated scanning method easily reduced the original 40,000 adjacent elements (0.075 mm 3 0.075 mm) down to a few ''elements of interest'' containing the Giardia cysts. We achieved an averaged signal-to-background ratio of 41.2 (minimum ratio of 12.1). Such high contrasts ensured the accurate mapping of all the potential Giardia cysts free of false positives or negatives. This was confirmed by the automatic retrieving and time-gated luminescence bioimaging of these Giardia cysts. Such automated microscopy based on time-gated scanning can provide novel solutions for quantitative diagnostics in advanced biological, environmental, and medical sciences. ' 2011 International Society for Advancement of Cytometry

show abstract

mentioning

confidence: 99%

Automated detection of rare‐event pathogens through time‐gated luminescence scanning microscopy

Lu¹,

Jin²,

Leif³

et al. 2011

Cytometry Pt A

View full text Add to dashboard Cite

show abstract

“…In recent years, flow cytometry has been gaining in popularity as a novel method of detecting and enumerating different parasites as Giardia cysts and Cryptosporidium oocysts present in environmental and fecal samples (Ferrari, 2003;Moss, 2001;Power, 2003). Many papers reported flow cytometry as a method more sensitive than either conventional or immunofluorescence microscopy for the detection of Giardia sp.…”

Section: Jacobbergermentioning

confidence: 99%

“…Many papers reported flow cytometry as a method more sensitive than either conventional or immunofluorescence microscopy for the detection of Giardia sp. cysts in fecal samples (Dixon, 1997;Dixon, 2002;Ferrari, 2003), in detection of Cryptosporidium in SCID mice (Arrowood, 1995), seeded horse feces (Cole, 1999), and seeded human stool specimens (Valdez, 1997). In addition to detection and enumeration, large-scale sorting could also be used in conjunction with flow cytometry to yield partially purified oocysts for research purposes, such as food-spiking and recovery experiments, viability determination, or molecular characterization (Dixon, 2005).…”

Section: Jacobbergermentioning

confidence: 99%

Applications of Flow Cytometry to Clinical Microbiology

Pieretti¹,

Masucci²,

Moretti³

2012

Clinical Flow Cytometry - Emerging Applications

View full text Add to dashboard Cite

“…These include intrinsic resistance to enzymatic degradation (they are not natural substrates for either nucleases or proteases), faster hybridization kinetics, higher binding affinities for their targets, and the ability to penetrate "difficult" biological structures such as the exosporium of freshly-germinated bacterial endospores, the cysts of parasites such as Cryptosporidium and Giardia (Jens Hyldig-Nielsen, personal communication) and the thick cell walls of bacteria such as Mycobacteriumand Listeria spp. (49,75,76). An additional advantage of PNA probes is their capacity for binding to portions of the ribosome that are physically inaccessible to traditional DNA probes, enabling detection of organism-specific diagnostic sequences that are otherwise "buried" in the higher-order structure of the ribosome (49,50,74,77).…”

Section: Biomimeticsmentioning

confidence: 99%

Methods for Whole Cell Detection of Microorganisms

Brehm-Stecher

2008

ACS Symposium Series

View full text Add to dashboard Cite

Microbes are ubiquitous, and can be found occupying nearly every imaginable niche on Earth. These include organic and inorganic surfaces, interfacial boundaries and within macroscopically solid matrices, such as the pore space of rocks. Because phylogenetically divergent microbes may be visually indistinguishable, understanding the species distribution and ecological significance of environmental microbes requires diagnostic tools that extend beyond simple phenotypic description. Methods for microbial diagnostics can be divided into two broad categories: cellular and acellular. Acellular techniques, such as the polymerase chain reaction or certain immunoassay formats may be effective at detecting molecular, structural or biochemical targets associated with specific cell types, but this information is provided out of its "natural", and arguably most meaningful context -that of the individual microbial cell. In contrast, cellular methods have the potential to preserve an abundance of valuable information. Apart from molecular identity, this includes information regarding cell morphology and other physiological characteristics, cell number and distribution within a sample, and physical or spatial associations with other cell types. The aim of this chapter is to provide an overview of whole cell methods for microbial detection, including both existing approaches and those still in development. The tools described here are expected to find wide application for the detection of microbes on surfaces or within complex matrices across a number of parallel or allied fields, including environmental, food and clinical microbiologies.

show abstract

Analysis‐only detection of Giardia by combining immunomagnetic separation and two‐color flow cytometry

Cited by 21 publications

References 26 publications

Automated detection of rare‐event pathogens through time‐gated luminescence scanning microscopy

Automated detection of rare‐event pathogens through time‐gated luminescence scanning microscopy

Applications of Flow Cytometry to Clinical Microbiology

Methods for Whole Cell Detection of Microorganisms

Contact Info

Product

Resources

About