Improved detection of viruses by electron microscopy after direct ultracentrifuge preparation of specimens

Hammond, Gregory; Hazelton, Paul R.; Chuang, Ilin; Klisko, Brian

doi:10.1128/jcm.14.2.210-221.1981

Cited by 64 publications

(29 citation statements)

References 11 publications

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“…By resuspending the pellet in a small volume, the virus is concentrated 50 -100fold. Alternatively, particles may be sedimented directly onto the grid using the Airfuge ® technique, resulting in similar enrichment factors (Gelderblom and Reupke, 1978;Hammond et al, 1981).…”

Section: Methods Used In Diagnostic Emmentioning

confidence: 99%

Diagnostic electron microscopy is still a timely and rewarding method

Biel

Gelderblom

1999

Journal of Clinical Virology

116

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In the history of diagnostic EM in virology, a change in use has been seen. Starting in the 1990s and coincident with the broad introduction of 'modern' diagnostic techniques, the number of EM diagnostic labs has decreased considerably--in spite of the obvious advantages of this technique. To guarantee the continuing performance of diagnostic EM in the future. EQA runs have to be performed as with other techniques in the diagnostic armament. The growing number of participants and participating countries indicates an interest in as well as a need for this program.

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Section: Methods Used In Diagnostic Emmentioning

confidence: 99%

Diagnostic electron microscopy is still a timely and rewarding method

Biel

Gelderblom

1999

Journal of Clinical Virology

116

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“…The enrichment factor for poxviruses is similar to that given by other authors using the airfuge (Müller et al 1981;Biel and Gelderblom 1999a), although it is difficult to make direct comparisons because the airfuge parameters were not identical and, moreover, enrichment factors were not determined at the detection limit. Higher enrichment factors might be achieved using the particle counting EM-90 Beckman rotor (Hammond et al 1981) although a direct comparison of the two rotor systems, including a critical evaluation of practical handiness, has yet to be published. Differences between the detection limit of spores and virus using airfuge enrichment may be explained by their different sizes and therefore, different susceptibilities to the mechanical forces exerted on the particles.…”

Section: Discussionmentioning

confidence: 99%

Detection limit of negative staining electron microscopy for the diagnosis of bioterrorism-related micro-organisms

Laue

Bannert

2010

Journal of Applied Microbiology

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Aims: To determine the detection limit of diagnostic negative staining electron microscopy for the diagnosis of pathogens that could be used for bioterrorism. Methods and Results: Suspensions of vaccinia poxvirus and endospores of Bacillus subtilis were used at defined concentrations as a model for poxviruses and spores of anthrax (Bacillus anthracis), both of which are pathogens that could be used for bioterrorist attacks. Negative staining electron microscopy was performed directly or after sedimentation of these suspensions on to the sample supports using airfuge ultracentrifugation. For both virus and spores, the detection limit using direct adsorption of a 10‐μl sample volume onto the sample support was 106 particles per ml. Using airfuge ultracentrifugation with a sample volume of 80 μl, the detection limit could be reduced to 105 particles per ml for spores and to 5 × 104 particles per ml for poxviruses. The influence on particle detection of incubation time, washing and adsorption procedures was investigated. Conclusions: The reproducibility and sensitivity of the method were acceptable, particularly considering the small sample volume and low particle number applied onto the sample support. Significance and Impact of the Study: Diagnostic negative staining electron microscopy is used for the diagnosis of pathogens in emergency situations because it allows a rapid examination of all particulate matter down to the nanometre scale. This study provides precise detection limit for the method, an important factor for the validation and improvement of the technique.

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“…All specimens were examined by negative-stain EM after concentration by direct ultracentrifugation (Hammond et al, 1981) and identified by specific morphology. An EIA rotavirus (Rotaclove, Cambridge Biosciences) was used on a seasonal basis from December to March.…”

Section: Electron Microscopymentioning

confidence: 99%

“…This study was undertaken to examine the individual adenovirus types from fecal specimens in children under 3 years of age in Manitoba, Canada, and to compare the methods of detection by virus culture (VC), EM with concentration by direct ultracentrifugation (Hammond et al, 1981), and EIA (Cambridge Bioscience). During July 1990-1991, 1187 fecal samples from children K3 years of age were collected at clinics and hospitals and submitted to the Cadham Provincial Laboratory, Winnipeg, Manitoba, Canada, for the diagnosis of virus infection.…”

Section: Introductionmentioning

confidence: 99%

Comparison of detection methods for adenovirus from enteric clinical specimens

Ahluwalia

Scott-Taylor

Klisko

et al. 1994

Diagnostic Microbiology and Infectious Disease

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Fecal samples submitted for virus examination over July 1990 to June 1991 from children < 3 years of age were examined by electron microscopy (EM), virus culture (VC), and enzyme immunoassay [EIA, group-reactive and adenovirus (Ad) 40/41 specific; Cambridge BioScience] to compare the detection rate of adenovirus from pediatric fecal specimens. Ad isolates of serotypes 1-7 grown in HEp-2 or primary rhesus monkey kidney cells were identified by neutralization. Graham 293 cell cultures were used only when specimens were found to be positive for Ad by EM, type-specific Ad40/41 EIA, and for isolates not identified by neutralization. Ads grown in 293 cells were identified by DNA restriction endonuclease analysis. Of the 1187 specimens examined, 105 (9%) were found to be positive for Ad. VC detected 93, while 12 additional positives were detected by EM or EIA. The relative sensitivity of VC, EIA, and EM for the 105 specimens was 89% (93), 45% (47), and 35% (37), respectively. Among the 105 positive specimens, enteric Ad, nonenteric Ad, and untypeable Ad were 28% (29), 65% (68), and 7% (8), respectively. Of 37 EM positives, 62% (23) were enteric Ad; 27% (10) were nonenteric including serotypes 2, 3, 4, 5, 12, and 31, with 4, 1, 1, 2, 1, and 1 isolates of each type positive, respectively; and 11% (4) were detectable only by EM. Five isolates were identified as variant of Ad 2(3), Ad 3(1) and Ad 31(1). Over a 1-year period, a single Ad41 variant strain was the most frequently detected enteric Ad in Winnipeg, Manitoba, Canada.(ABSTRACT TRUNCATED AT 250 WORDS)

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Improved detection of viruses by electron microscopy after direct ultracentrifuge preparation of specimens

Cited by 64 publications

References 11 publications

Diagnostic electron microscopy is still a timely and rewarding method

Diagnostic electron microscopy is still a timely and rewarding method

Detection limit of negative staining electron microscopy for the diagnosis of bioterrorism-related micro-organisms

Comparison of detection methods for adenovirus from enteric clinical specimens

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