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PubMed2003

Practically relevant research: capturing real world tasks, environments, and outcomes.

PURPOSE: Development of strategies to optimize the functional performance of older adults requires understanding the behavior of older people doing tasks in real-world settings and capturing these interactions in research protocols. This is a major challenge as there is some degree of tension between capturing the contextual variables and constraints that operate in the real world and the scale of research that can be realistically conducted within controlled experimental settings. This article presents a research approach that can be used to help ensure the ecological validity of research protocols. The intent is to demonstrate how an ecologically valid approach affords greater insight into the performance of older adults in real world settings. METHODS: The approach involves techniques such as task analysis and simulation. Examples from two research projects examining aging and the performance of real-world computer-based work tasks are used to demonstrate the application of this approach. RESULTS: The article demonstrates how an ecologically valid research approach yields information about human performance that can be translated into solutions for real-world problems. IMPLICATIONS: Ecologically valid research protocols offer the potential for providing answers to real-world problems and advancing theory regarding aging and functional performance.

Activities of Daily Living↗

PubMed1996

Isolation of genes amplified in human cancers by microdissection mediated cDNA capture.

It has been increasingly recognized that homogeneously staining regions (hsr) in human cancers may be complex structures composed of large amplified DNA domains containing multiple genes. It is therefore important to devise strategies for the rapid isolation of cDNAs expressed from these structures. Using a procedure we term microdissection mediated cDNA capture, we recovered hsr specific cDNAs from two different human tumors. The glioblastoma cell line TX3868 and the human sarcoma cell line OsA-CL carry hsrs containing amplified sequences from chromosome 12q13-15. We recovered 17 hsr specific cDNAs following microdissection of these hsrs which had been previously hybridized in situ with linkered cDNA. Northern blot analysis with these cDNAs revealed hybridization to distinct transcripts in OsA-CL RNA and TX3868 RNA. None of the OsA-CL cDNA clones showed cross hybridization with the TX3868 cDNAs suggesting that despite their coincident band localization on 12q, the OsA-CL and TX3868 amplification units do not completely overlap. These results significantly increase the number of amplified genes assigned to the 12q13-15 amplicon illustrating both the complexity of hsrs derived from this region and the utility of microdissection mediated cDNA capture to gain rapid access to cDNAs transcribed from amplified genes.

Base Sequence↗

PubMed2000

Record-linkage between two anonymous databases for a capture-recapture estimation of underreporting of AIDS cases: France 1990-1993. The Clinical Epidemiology Group from Centres d'Information et de Soins de l'Immunodéficience Humaine.

OBJECTIVE: To estimate the completeness of the French mandatory AIDS surveillance system (Declaration Obligatoire DO) over the 1990-1993 period using a capture-recapture approach, by matching the mandatory reports with the AIDS cases present in the French Hospital Database on HIV infection (FHDH). METHODS: An anonymous record-linkage algorithm was developed to identify those cases common to both anonymous surveillance systems. The linkage was based on sex, date of birth, and infection risk group, all strictly matched, and on the dates of AIDS diagnosis and of death, the places of diagnosis and residence, and the AIDS-defining diseases at diagnosis. The total number of AIDS cases and completeness of both surveillance systems were estimated using a capture-recapture approach, assuming independence of the ascertainment sources. RESULTS: The completeness of the mandatory reporting was estimated at 83.6% (95% CI: 82.9-84.3), and that of the FHDH at 47.6% (95% CI: 46.9-48.3) for the surveillance of AIDS cases diagnosed among adults in France between 1990 and 1993. The completeness of the system based on FHDH increased over the study period as more hospitals joined the project, while the completeness of the DO surveillance system remained stable. CONCLUSION: This approach was useful in estimating the underreporting of AIDS cases in France. Regularly performed, it will allow the impact of underreporting to be monitored over time.

Acquired Immunodeficiency Syndrome↗

PubMed1988

Serodiagnosis of early Lyme disease: analysis of IgM and IgG antibody responses by using an antibody-capture enzyme immunoassay.

We used an antibody-capture enzyme immunoassay (EIA) to evaluate the early antibody responses to Borrelia burgdorferi in paired sera from 30 patients with erythema chronicum migrans. During acute disease, 20 (67%) patients had elevated specific IgM responses, and by convalescence (one to four weeks after treatment), 28 (93%) patients had increased IgM or IgG responses. In acute specimens, elevated IgM responses correlated with disseminated infection; however, by convalescence, most patients with either localized or disseminated disease had positive tests. Among 133 control subjects, IgM cross-reactivity was observed in 4 of 37 patients with either Epstein-Barr virus or rickettsial infections, and false-positive IgG tests were seen in 8 of 28 patients with syphilis. With antibody-capture EIA, the diagnosis of Lyme disease can be confirmed in the majority of acutely ill patients and in almost all patients by convalescence.

Acute Disease↗

PubMed1996

Comparative detection of measles-specific IgM in oral fluid and serum from children by an antibody-capture IgM EIA.

In vaccinated populations, the diagnosis of measles often requires laboratory confirmation. Serum tested by EIAs has proven sensitive and specific for diagnosing measles. For comparison of detection of measles-specific IgM in oral fluid and serum samples by an antibody-capture EIA, 163 Ethiopian infants who presented for routine measles vaccination were studied. Paired serum and oral fluid samples were collected before and 2 weeks after vaccination; 269 paired samples were adequate for analyses. Of the 104 serum samples that were IgM-positive, 95 (91%) of the paired oral fluid samples were IgM-positive. Of the 165 serum samples that were IgM-negative, 156 (95%) of the paired oral fluid samples were IgM-negative. The Pearson partial correlation coefficient for optical density readings from postvaccination oral fluid compared with serum was 0.81. Oral fluid appears to be an acceptable alternative to serum for measuring measles-specific IgM antibodies by an antibody-capture EIA.

Antibodies, Viral↗

PubMed1997

Diagnosis of measles with an IgM capture EIA: the optimal timing of specimen collection after rash onset.

The optimal timing for collection of a single serum specimen to diagnose measles by using a monoclonal antibody-capture EIA was evaluated. Results of testing paired serum samples from 166 measles cases with at least 1 IgM-positive specimen were analyzed. Among persons whose second samples were IgM-positive, the seropositivity rate for first samples was 77% when collected within 72 h and 100% when collected 4-11 days after rash onset. Among unvaccinated persons whose first samples were IgM-positive, the rate for IgM positivity of second specimens declined from 100% at 4 days to 94% at 4 weeks after rash onset, then declined further to 63% at 5 weeks. Some previously vaccinated persons became IgM-negative during the third week after rash onset. In general, a single serum specimen collected between 72 h and 4 weeks after rash onset can be used to diagnose most cases of measles with an IgM capture EIA.

Adolescent↗

PubMedDate not supplied

Measurement of clonazepam by electron-capture gas-liquid chromatography with application to single-dose pharmacokinetics.

A simple, sensitive, and rapid method for analysis of clonazepam in blood by gas-liquid chromatography with electron-capture detection is described. The method employs neutral benzene extraction from plasma and concentration into a small volume for autoinjection into the electron capture detector-equipped chromatograph. This method consistently provides blank plasma samples free of contaminants in the areas corresponding to the retention times for clonazepam and desmethylflunitrazepam (the internal standard). Sensitivity is 1 ng/mL, and standard curves are linear from 1 to 200 ng/mL. The applicability of the method to single-dose pharmacokinetics is illustrated.

Adult↗

PubMed2003

Evaluation of two prototype traps and existing trap designs for captures of Japanese beetles (Coleoptera: Scarabaeidae).

Two prototype trap designs and four other trap designs currently on the market were evaluated for capture of Japanese beetles, Popillia japonica Newman. The standard Japanese beetle trap manufactured by Trece (Palo Alto, CA) and a prototype trap captured consistently more beetles than other trap designs tested. Traps that used a bag for collection of beetles were not as effective because beetles were able to escape through the holes made for drainage. The new trap design allows traps to be transported more easily and stored in a smaller area because of the collapsible design. This is a significant improvement for states that use thousands of traps for monitoring Japanese beetles.

Animals↗

PubMed1991

Funnel trap for the capture of phlebotomine sand flies (Diptera: Psychodidae) from tree holes.

A funnel trap that fitted over holes leading into hollow trees was used to capture adult phlebotomine sand flies, Lutzomyia shannoni Dyar, on Ossabaw Island, Chatham County, Ga. These insects rested in hollow trees during the day and were collected by funnel traps as they egressed from the tree holes at night. The trap is lightweight, durable, inexpensive, waterproof, and selective. Using this trap, greater than 100 healthy L. shannoni ++were captured per night by a single investigator during July and August 1988 when adult flies were abundant on the island.

Animals↗

PubMed1993

Mosquitoes (Diptera: Culicidae) captured in the Iquitos area of Peru.

A mosquito capture program was initiated to study mosquito species and their potential for arboviral transmission in the Peruvian Amazon. More than 35,000 mosquitoes of 13 different genera and at least 25 species were captured in urban and sylvan sites in the Iquitos area. These findings represent the first published list of Peruvian mosquitoes since 1971 and the first such list from the Peruvian Amazon.

Animals↗

PubMed1990

Branch capture reactions: effect of recipient structure.

Branch capture reactions (BCR) contain two DNA species: (i) a recipient restriction fragment terminating in an overhang and (ii) a displacer-linker duplex terminating in a displacer tail complementary to the overhang as well as contiguous nucleotides within the recipient duplex. Branched complexes containing both species are captured by ligation of the linker to the recipient overhang. Specificity depends upon branch migration and is increased by substitution of bromodeoxycytidine for deoxycytidine in the displacer. BCR rates and specificities were determined for recipient overhangs that were (i) 5' and 3', (ii) 3 and 4 nucleotides long, and (iii) 0-100% G+C. Model systems permitted independent determination of G+C and branching effects on ligation rates and verification of rapid equilibrium between the branched complex and its component species. With all 4-base overhangs, recipient duplexes permitting extensive branch migration became saturated with displacer-linker duplexes. With increasing G+C, increasing ligation at competing sites led to decreased BCR specificity. BCR may be used to label a DNA fragment prior to electrophoresis, mark a fragment for affinity chromatography, or introduce a new overhang sequence compatible with a restriction endonuclease site in a cloning vector. A protocol was confirmed for mapping restriction sites in cloned DNA.

Base Sequence↗

PubMed1991

Branch capture reactions: displacers derived from asymmetric PCR.

Branch capture reactions (BCR) contain three DNA species: (i) a recipient restriction fragment terminating in an overhang, (ii) a displacer strand containing two adjacent sequences, with one complementary to the overhang and to contiguous nucleotides within the recipient duplex and (iii) a linker which is complementary to the second displacer sequence. Branched complexes containing all three species may be captured by ligation of the linker to the recipient overhang. The use of 5-MedC in the displacer facilitates BCR. High temperature ligation with a thermostable enzyme increased specificity for ligation to the correct recipient in a complex mixture of restriction fragments. Displacer synthesis by PCR permitted separate reactions of formation of stable displacement complexes and of high-temperature ligation. Ethylene glycol-containing buffer permitted PCR with 5-MedCTP or high G + C products using thermostable polymerases. BCR may be used to modify the ends of one recipient DNA duplex in a population of duplex DNA fragments. Modification of the recipient could be used to facilitate detection, affinity chromatography or cloning. By using PCR to obtain a BCR displacer, the sequence non-homologous to the recipient duplex may be expanded to include the sequence of a selectable marker, thus facilitating chromosome walking.

Base Sequence↗

PubMed1993

Microtiter format gene quantification by covalent capture of competitive PCR products: application to HIV-1 detection.

We have developed a simple gene quantification system using the competitive polymerase chain reaction (CPCR) followed by microtiter format analysis. CPCR is carried out using a mutant competitor with the same size as the target DNA product, and a minimal base exchange to insure the same amplification kinetics. One primer is aminated at the 5' end to produce PCR products that are captured onto carboxylated wells of microtiter plates through peptide bond formation. The non-aminated DNA strands are stripped off from the wells by alkali washing, and the remaining aminated strands are hybridized with either a digoxigenin-labeled wild type-specific oligonucleotide probe or a competitor-specific probe. To standardize the hybridization conditions of the probes, a DNA construct containing wild type and mutant competitor sequences in tandem is captured at different concentrations, hybridized with the probes, and used to generate a standard curve. Bound probes are detected by anti-digoxigenin antibody conjugated with peroxidase and chromogen. Optical densities are recorded with a conventional microtiter plate reader and converted to concentrations according to the standard curves. The ratios of wild type DNA to mutant competitor are used to determine the initial amounts of wild type DNA in the samples. This method was used successfully to quantify human immunodeficiency virus type 1 (HIV-1) env gene in human lymphocytes. It only requires a thermal cycler and a conventional microtiter plate reader, and can be readily done on a large scale. Potential applications include detection of other pathogens, diagnosis of genetic disorders and studies of gene expression.

Base Sequence↗

PubMed1997

Enrichment of oligo(dG).oligo(dC)-containing fragments from human genomic DNA by Mg 2+-dependent triplex affinity capture.

Oligo(dG).oligo(dC)- or short poly(dG).poly(dC)-containing fragments were enriched and cloned by means of Mg2+-dependent triplex affinity capture and subsequent cloning procedures. A library constructed after three cycles of enrichment showed that approximately 80% of the clones in the supercoiled form formed a complex with labeled oligonucleotide (dG)34. However, while the rest of the clones retained the ability to form a complex (type I clones), 90.9% failed to form a complex when they were linearized. This group of DNA was abundant in the genomic DNA, although it showed only approximately 3-fold enrichment by one cycle of affinity capture. This group was further classified into two species (types II and III) based on complex formation ability after phenol extraction. Type II clones retained the complex formation ability after treatment, while the human telomere [(TTAGGG)n] and telomere-like [(TGGAA)n] or [(TGGAG)n] sequences belonging to type III clones did not. Serial deletion experiments and the binding assays using oligonucleotides confirmed that the repetitive units containing T(G)nT ( n = 3-5) tracts or (G)n-motifs (n >/= 3) were the sites of complex formation for type II and III clones. On the other hand, type I clones contained poly(dG).poly(dC) tracts at least 10 nt long, and DNase I-footprinting analysis indicated that these tracts were the sites of complex formation.

Cloning, Molecular↗

PubMed1999

Random mutagenesis by recombinational capture of PCR products in Bacillus subtilis and Acinetobacter calcoaceticus.

We describe a general method for random mutagenesis of cloned genes by error-prone PCR or DNA shuffling that eliminates the need for post-amplification subcloning following each cycle of mutagenesis. This method exploits the highly efficient and recombinogenic nature of DNA uptake during natural transformation in the Gram-positive bacterium Bacillus subtilis and the Gram-negative bacterium Acinetobacter calcoaceticus. Plasmid systems were designed that allow capture of PCR-amplified DNA fragments by marker-replacement recombination with a structurally similar helper plasmid resident in the transformation recipient. This recombination event simultaneously transfers the amplified sequences into the helper plasmid and restores the integrity of a drug resistance gene, thereby affording a direct selection for fragment capture. Although this strategy was sufficiently effective to permit recovery in B. subtilis of up to 10(3) transformants/microgram of PCR product, equivalent plasmid systems were approximately 100 times more efficient in A.calcoaceticus. Acinetobacter calcoaceticus also offers the advantage of essentially constitutive transformation competence in ordinary complex broth, such as LB, in contrast to two-step growth in semi-synthetic media required for optimal transformation of B.subtilis.

Acinetobacter calcoaceticus↗

PubMed2000

Mass spectrometry of single-stranded restriction fragments captured by an undigested complementary sequence.

In this report, we describe a simple and accurate method to analyze restriction fragments using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. The two complementary strands of restriction fragments are separated through hybridization to a capture probe, which is a single-stranded undigested fragment. Using the biotin-streptavidin linkage, the hybrid is immobilized on streptavidin-coated magnetic beads. After conditioning the captured restriction fragments, they are eluted from the probe and their molecular weights are determined. The proposed method greatly improves the quality, and reduces the complexity of the mass spectrum by analyzing only one of the complementary strands of restriction fragments.

Biotinylation↗

PubMed2005

Oligonucleotide-assisted cleavage and ligation: a novel directional DNA cloning technology to capture cDNAs. Application in the construction of a human immune antibody phage-display library.

The use of oligonucleotide-assisted cleavage and ligation (ONCL), a novel approach to the capture of gene repertoires, in the construction of a phage-display immune antibody library is described. ONCL begins with rapid amplification of cDNA ends to amplify all members equally. A single, specific cut near 5' and/or 3' end of each gene fragment (in single stranded form) is facilitated by hybridization with an appropriate oligonucleotide adapter. Directional cloning of targeted DNA is accomplished by ligation of a partially duplex DNA molecule (containing suitable restriction sites) and amplification with primers in constant regions. To demonstrate utility and reliability of ONCL, a human antibody repertoire was cloned from IgG mRNA extracted from human B-lymphocytes engrafted in Trimera mice. These mice were transplanted with peripheral blood lymphocytes from Candida albicans infected individuals and subsequently immunized with C.albicans glyceraldehyde-3-phosphate dehydrogenase (GAPDH). DNA sequencing showed that ONCL resulted in efficient capture of gene repertoires. Indeed, full representation of all V(H) families/segments was observed showing that ONCL did not introduce cloning biases for or against any V(H) family. We validated the efficiency of ONCL by creating a functional Fab phage-display library with a size of 3.3 x 10(10) and by selecting five unique Fabs against GAPDH antigen.

Adolescent↗

PubMed1984

Application of a capture-recapture method (the Bernoulli census) to historical epidemiology.

Obstacles to historical epidemiologic research are formidable, especially for periods prior to the 19th century. Linguistic and paleographic difficulties are compounded by haphazard contemporary documentation and subsequent archival wastage, making real variations in disease frequency difficult to distinguish from record loss. This report investigates the application of a capture-recapture model, adapted from work on mobile animal populations, to this problem or archival loss. With the use of this technique, the original size of a psychiatric case population is estimated from incomplete extant enumerations. The cases are derived fron the Court of Wards and Liveries (1540-1660), the first statutory institution in England responsible for protecting persons legally incompetent due to mental disability. Documentation on this population survives in at least 10 record collections, the most extensive containing at best 82% of all original cases, the least extensive, 1%. Calculations based on extant files indicate a fourfold increase in the incidence of mental disability certifications between 1540-1549 and 1630-1639. The results of applying the Bernoulli census method to these materials eliminate biased archival loss as a plausible explanation for this rise. The origin of these changes in altered court policy or procedure, population growth, or increasing rates of mental disability awaits further investigation. While the Bernoulli census computations with historical data are mathematically equivalent ot those employed in recent epidemiologic applications, this specific data set prompts a conceptual revision of the Bernoulli model from capture to loss of random samples.

Animals↗