Diagnostic value of telescoping plugged catheters in mechanically ventilated patients with bacterial pneumonia using the Metras catheter

A new guiding technique, Metras catheter (MC), for blindly introducing a telescoping plugged catheter (TPC) was applied to 25 mechanically ventilated patients with suspected bacterial pneumonia (BPN). Results obtained with TPC-MC were compared with those obtained with TPC using a conventional fiberoptic bronchoscope (FB) in random order. The diagnosis of BPN was definitely confirmed in 18 patients. In 7 patients, all TPC samples (MC and FB) were sterile, and a diagnosis other than BPN was proved. In the former group, colony-forming units equal to or greater than 10(3)/ml of one or more microorganisms were obtained in 61% of TPC-MC and in 66% of TPC-FB samples. These percentages increased to 64 and 71%, respectively, when 4 patients with previous antibiotic treatment were excluded from the study group. Agreement was observed between microorganisms cultured from both TPC samples in 11 of 18 patients with proved BPN (61%). Complete disparity was seen only in 2 patients (11%). Two patients developed a self-limiting hemoptysis after the TPC procedure (MC and FB, respectively). We conclude that TPC-MC is both a sensitive and specific technique for the diagnosis of BPN in mechanically ventilated patients. Because the diagnostic value of TPC-MC is similar to that of TPC-FB, we propose that the MC be used in patients receiving mechanical ventilation when the FB is not available. The simplicity and lower cost of this new system are important advantages to be considered over the fiberoptic bronchoscope.     

Rapid detection of anticardiolipin antibodies

A rapid screening method for the detection of antiphospholipid antibodies is described. Dense, red dyed polystyrene beads coated with cardiolipin were incubated with test sera for a short period of time, then added to a microtube containing anti-human IgG in a gel provided within a pre-cast card (DiaMed ID Microtyping System). The card was centrifuged at 150g for 5 min and then examined for movement of the beads through the gel. Beads without bound antibody travelled through the gel and formed a pellet on the bottom of the tube. Anti-human IgG within the gel matrix impeded cardiolipin-coated beads when antiphospholipid antibodies bound to the beads. Positivity was indicated by the formation of a layer of beads on the top of the gel matrix. Prospective analysis of 103 samples for the presence of antiphospholipid antibodies by flow cytometry and the gel-card technique showed good correlation between the two methods. All samples found to be positive by flow cytometry (23 of 103) were identified as positive by the gel-card technique. Two samples were identified as positive by the gel-card method but negative by flow cytometry. The technique is simple to perform and should prove useful as a rapid screening method for the detection of antiphospholipid antibodies. Am. J. Hematol. 57:315–319, 1998. © 1998 Wiley-Liss, Inc.     

Using wild white-tailed deer to detect eastern equine encephalitis virus activity in Maine

Serum from 226 free-ranging white-tailed deer (Odocoileus virginianus) was screened for Eastern Equine Encephalitis Virus (EEEV) antibodies using plaque reduction neutralization tests. EEEV antibodies were detected in 7.1% of samples. This is the first time EEEV antibodies have been detected in O. virginianus populations in the state of Maine (ME). The highest percentage of EEEV positive sera was in Somerset County (19%) in central ME, and this is the first time that EEEV activity has been detected in that County. EEEV RNA was not detected in any of the 150 harvested deer brain samples submitted to the ME Department of Inland Fisheries and Wildlife as a part of screening for Chronic Wasting Disease. This suggests that screening deer brains is not an efficient method to detect EEEV activity. For each serum sample tested, the geographic location in which the deer was harvested was recorded. Significant spatial clustering of antibody-positive sera samples was not detected. Relative to seronegative deer, seropositive deer were slightly more likely to be harvested in nonforested areas compared with forested areas. Results indicate that screening of free-ranging deer sera can be a useful tool for detecting EEEV activity in ME and other parts of North America.     

An Antibody Screening Test Based on the Antiglobulin Gel Technique, Pooled Test Cells, and Plasma

The recently introduced gel technique offers significant advances compared to traditional tube tests. The purpose of the present study was to design an antibody screening test based on the gel technique, pooled cells, and plasma and to evaluate this test by comparison with a conventional spin-tube indirect antiglobulin test (IAT) combined with a two-stage papain technique. Pilot studies were performed to establish optimal parameters during the design phase, and finally 5,446 consecutive samples were screened for irregular antibodies by the gel technique in parallel with routine techniques. Irregular erythrocyte antibodies were detected in 151 samples, and the gel technique proved equal or superior to the tube test in the detection of all antibodies except 'enzyme-only' anti-E and anti-Le^a. We conclude from this study that screening for unexpected antibodies using the gel IAT in our set-up, which includes: (1) omission of enzyme technique; (2) the use of stabilized (EDTA) plasma instead of serum as test material in order to facilitate automation; and (3) pooling 2 by 2 of 4 test cells (to make the gel technique price competitive), is a fast, reliable and sensitive procedure that maintains transfusion safety and compares favourably with our previous routine of saline IAT combined with a two-stage papain technique.     

Rapid method for the detection of anti-HCV antibodies in patients with chronic hepatitis C.

OBJECTIVE: to evaluate the usefulness of a simple, rapid, qualitative technique (MedMira Rapid Test) to detect antibodies against hepatitis C virus (HCV) and compare this approach with an immunometric technique in patients with chronic hepatitis C infected with different genotypes. METHODS: anti-HCV antibodies were determined with the MedMira rapid technique and an immunometric method in samples from 138 patients with chronic hepatitis C infected with different HCV genotypes, and in 50 samples from healthy individuals. RESULTS: the MedMira rapid technique detected anti-HCV antibodies in 135 (98%) of 138 serum samples from patients with chronic hepatitis C, whereas the immunometric method gave positive results in all 138 samples. Three of the 138 anti-HCV-positive samples identified with the immunometric method and confirmed by inmunoblotting were repeatedly negative with the MedMira rapid technique. Two of these samples were genotype 1 and the third was not genotyped. All samples from the control group were negative for anti-HCV antibodies by both methods. The sensitivity and specificity of the MedMira rapid technique relative to the immunometric technique were 98% and 100% respectively. CONCLUSION: the MedMira rapid technique is a quick, specific and sensitive method that is easy to use by nonspecialized personnel, and is a good alternative to other, slower methods for the diagnosis of chronic hepatitis C.     

Two-stage papain-indirect antiglobulin test and LISS direct agglutination are not appropriate for pretransfusion screening for unexpected antibodies.

BACKGROUND. The screening for unexpected antibodies has proved to be a suitable pretransfusion compatibility method, but controversy still remains regarding the most appropriate serologic techniques to use in such a screening and, in particular, whether or not to use enzyme-based methods. METHODS. Over a period of three years, 27,149 patient sera submitted to pretransfusion testing were screened for unexpected antibodies. Serologic techniques included LISS-direct agglutination (DAG) reading plus indirect antiglobulin test (IAT), and two-stage papain (2SP)-IAT. RESULTS. In all, 592 (2.18%) serum samples yielded a positive result. Further studies of these specimens disclosed 466 alloantibodies in 371 cases, and 221 unwanted positive reactivities. 2SP-IAT and LISS-DAG allowed the detection of 124 alloantibodies missed by LISS-IAT (78 anti-Lewis, 33 anti-E, 7 anti-P1, 2 anti-K1, 2 anti-K3, 1 anti-M, 1 anti-Cw), but were responsible for 81% of the unwanted positive reactivities. CONCLUSIONS. Since most alloantibodies detected only by 2SP-IAT or LISS-DAG were of doubtful clinical significance, and these techniques produced a high number of unwanted positive reactivities, we conclude that 2SP-IAT and LISS-DAG are not appropriate for the pretransfusion screening for unexpected antibodies.     

Evaluation of an enzyme immunoassay for the combined detection of antibodies to HIV-1, HIV-2, HTLV-I and HTLV-II.

OBJECTIVE: To assess the sensitivity and specificity of a newly-developed assay (Bioelisa HIV-1 + 2, HTLV-1 + 2) for the simultaneous detection of HIV-1, HIV-2, HTLV-I and HTLV-II antibodies in human serum or plasma specimens. METHODS: A panel of 775 well characterized serum or plasma samples was studied. This included samples confirmed to contain antibodies to HIV-1 (n = 46), HIV-2 (n = 19), HTLV-I (n = 49) and HTLV-II (n = 12), samples containing low titres of anti-HIV antibody (n = 14) and samples collected during HIV seroconversion (n = 36). Eighty-three sera samples which were reactive in one or more HIV or HTLV screening assays, but which could not be confirmed to contain anti-HIV-1/2 or anti-HTLV-I/II antibodies, were also examined. RESULTS: Excluding the seroconversion samples and those selected on the basis of false reactivity in other screening assays, the Bioelisa kit had a sensitivity of 100% for antibody to all four viruses and a specificity of 98.8%. The ability of the kit to detect anti-HIV during seroconversion was similar to that of several other synthetic HIV-antigen-based screening kits currently in use. CONCLUSIONS: Our findings indicate that the Bioelisa kit is sufficiently accurate to screen for both HIV and HTLV infections and that it warrants larger scale trials. Its use might allow blood donor screening for HTLV infection to be introduced more widely at modest extra cost [corrected].     

Diagnostic value of quantitative cultures of bronchoalveolar lavage and telescoping plugged catheters in mechanically ventilated patients with bacterial pneumonia

We compared the diagnostic value of quantitative cultures of bronchoalveolar lavage (BAL) and telescoping plugged catheter (TPC) samples in 34 nonimmunocompromised, mechanically ventilated (MV) patients with suspected bacterial pneumonia. A control group of seven "noninfected" MV patients was also studied. In 92% of patients with bacterial pneumonia (32 of 34), simple endotracheal aspiration samples recovered one or more microorganisms. Both BAL and TPC samples cultured colony-forming units (cfu) greater than or equal to 10(3)/ml of one or more microorganisms in 56% (19 of 34) of patients. TPC and BAL culture results agreed on 88.5% (54 of 61) of the recovered microorganisms. Sterile TPC and BAL cultures agreed on 80% (4 of 5) of the cases. Microorganisms cultured from blood samples were also cultured from BAL and TPC specimens. Culture results from the two techniques completely disagreed in only one case (3%). In the control group, one TPC and two BAL cultures yielded microorganisms in cfu greater than or equal to 10(3)/ml. Specificities of BAL and TPC were 71 and 86%, respectively, whereas specificity of endotracheal aspiration was only 14%. Both the bacterial index obtained by TPC and BAL, as well as the quantitative cultures, correlated moderately well (r = 0.78 and 0.72, respectively, p less than 0.001 for both correlations). BAL and TPC results caused changes of antibiotic treatment in 11 of 23 survivors. Neither BAL nor TPC caused complications. Our results demonstrate that BAL and TPC diagnose bacterial pneumonia in MV patients with similar accuracy. Culture results from both techniques showed excellent qualitative and reasonable quantitative agreement.     

5种不规则抗体筛查方法的检测阈值比较

目的:了解各种不规则抗体筛查方法的检测阈值,以避免不规则抗体漏检而引起输血不良反应。方法:选取献血者不规则抗体筛查阳性样本3例和配血不合疑难样本9例,采用盐水法、酶介质法、凝聚胺法、抗人球蛋白法、微柱凝胶法平行检测,比较5种方法的检测阈值。结果:盐水法只能检出IgM类的不规则抗体;木瓜酶法在检测IgG抗体效价时敏感性略低,而且有漏检的可能;凝聚胺法检测结果不易判断;抗人球蛋白法操作较繁琐;微柱凝胶法对IgG类抗体检测效价较高,高于凝聚胺法和抗人球蛋白法。结论:微柱凝胶法操作简单,检出率较高,是值得推广的不规则抗体筛查方法。     

Rapid-detection GBM-ANCA ELISA. An emergency tool for the early diagnosis of type I and II rapidly progressive glomerulonephritis

Rapidly progressive glomerulonephritides (RPGN) are forms of necrotizing glomerulonephritis associated with anti-glomerular basement membrane (anti-GBM) and anti-neutrophil cytoplasmic antibodies (ANCA) against the antigens proteinase-3 (anti-PR3) and myeloperoxidase (anti-MPO). RPGN have a course of rapid progression to renal failure. We compared the results from the semiquantitative ELISAs for anti-GMB antibodies, PR3-ANCA and MPO-ANCA and the indirect immunofluorescence technique (IIF) against a new rapid assay (30 minutes) for the same antibodies in patients with clinically suspected RPGN. The semiquantitative ELISAs for anti-GBM antibodies and PR3-ANCA and MPO-ANCA have a proven diagnostic significance in patients with RPGN I and III. There were no significant differences between the ANCA-GBM screening test and the results from the semiquantitative ELISAs (p > 0.05). We did not find significant differences between the results for PR3-ANCA and MPO-ANCA from the ANCA-GBM screening test with C-ANCA and P-ANCA IIF values (p > 0.05). We also corroborated that the ANCA-GBM screening test is a diagnostic tool for RPGN I and III as useful as the semiquantitative ELISAs and the IFF technique. The ANCA-GBM ELISA screening test is a tool as useful as the semiquantitative ELISA against anti-GBM antibodies for diagnosis of RPGN I. The comparison of the screening ELISA with the IIF technique and the semiquantitative ELISAs against PR3-ANCA and MPO-ANCA showed similar utility for diagnosis of RPGN III. The advantages of the new screening assay are that three antibodies are tested at the same time, yielding results in only 30 minutes.     

A Solid-Phase Test Using Antiglobulin and Enzyme-Enhanced Techniques for Erythrocyte Antibody Screening during Pregnancy

We have modified previously described solid-phase tests for erythrocyte antibody screening to develop a method, suitable for antiglobulin- and enzyme-enhanced techniques (SPH-IAT and SPH-ENZ). In this study we compared the SPH tests with an autoanalyzer (AA) technique. The results were more specific with the SPH tests than with the AA. Of 4,234 unselected samples from pregnant women, screen-positive samples were reduced from 96 (2.27%) by the AA, to 56 (1.32%) by the SPH tests. This difference was due to the reduced number of false-positive reactions with the SPH tests, 0.47% compared to 1.44% with the AA. Antibodies detected by the AA and the SPH-IAT and -ENZ were: 9 Rh prophylaxis, 2 anti-c, 1 anti-K and anti-M, and 1 anti-Jk^a. Antibodies detected only by the SPH tests were 1 anti-K, 1 anti-Le^a (SPH-IAT and -ENZ), 1 anti-M (SPH-IAT) and 4 anti-Jk^a (SPH-ENZ). One anti-C, 2 anti-D, 3 Rh prophylaxis and 1 anti-E were detected by the AA and the SPH-ENZ but failed to react by the SPH-IAT. One anti-Le^aand 8 Rh prophylaxis antibodies were detected by AA only. All clinically important antibodies were detected by the SPH tests. We conclude that the SPH-IAT and SPH-ENZ are screening methods with high specificity that are readily adaptable to larger series of samples from pregnant women and suitable for automated handling throughout the screening and identification process.     

catELISA: a facile general route to catalytic antibodies.

The low abundance and activity of catalytic antibodies are major obstacles to their selection from the virtually unlimited repertoire of antibody binding sites. The requirement for new screening methodologies is further emphasized by the availability of combinatorial libraries, in which a functional polypeptide has to be selected out of millions of possibilities. We present a simple and sensitive screening approach (termed catELISA) based on immobilized substrates and immunodetection of the end product of the catalyzed reaction. The feasibility of catELISA is demonstrated here by the generation of potent ester-hydrolyzing antibodies by direct screening of hybridoma supernatants. We show that this approach is not only facile but general: it is not limited by type of reaction, substrate, or catalyst (enzymes, catalytic antibodies, chemical catalysts). catELISA opens a route to catalytic antibodies that replaces existing lengthy and arduous methods, thus allowing us to expand their number and improve their quality and to address questions that would otherwise be difficult to answer.     

Detection of Irregular Red Cell Antibodies: More than 3 Years of Experience with a Gel Technique and Pooled Screening Cells

Background and objectives: The purpose of this study was to evaluate more than 3 years of experience with a gel technique in combination with pooled screening cells for the detection of irregular red cell antibodies. Materials and methods: Conventional serologic methods were used for blood typing, antibody screening and cross-matching until the end of 1992. We introduced the gel technique as a routine assay for antibody detection and identification in 1993. Results: After the tube technique had been abandoned, the number of false-positive antibody screening tests was reduced by 71%, positive antibody screening tests by 33%, enzyme agglutination by 100% and rouleaux reactions and cold-reacting antibodies by more than 50%. There was a 40% increase in first-time detection of clinically relevant antibodies. We saw no increase in delayed haemolytic transfusion reactions. Conclusions: For the detection of irregular red cell antibodies, pooled screening cells in combination with a gel technique are at least as efficient and safe as a conventional tube technique with unpooled test cells.     

Evaluation of a Solid-Phase Test for Erythrocyte Antibody Screening of Pregnant Women, Patients and Blood Donors

We have studied a previously described solid-phase test (SPH) for screening and identification of antibodies directed against erythrocyte antigens, in large sample numbers and throughout routine processing. The screening results of all samples from pregnant women, patients and blood donors (n = 36,701) sent to our blood bank from August 1992 to July 1993 were compiled and evaluated. Pregnant women were screened by the SPH-antiglobulin technique (IAT) and the SPH-enzyme-enhanced technique (ENZ), while the patients and blood donors were screened only by the SPH-IAT. Samples with known alloimmunization were excluded from this study. Positive screening reactions were further investigated by the SPH-IAT and SPH-ENZ techniques, the manual hemagglutination tests low ionic-strength solution IAT and 2-stage papain. Pregnant women: A positive reaction in one or both of the SPH tests was found in 1.2% of the samples. The SPH tests identified 99 (98.0%) and the manual methods 79 (78.2%) of a total of 101 antibodies. Of the identified antibodies, 61 were to antigens in the Rh blood group system, including 34 Rh immune globulin. The remainder had specificity directed against antigens K, Jk^a, M, S, Le^a and Le^b. Patients: A positive reaction in the SPH-IAT test occurred in 0.9% of the samples. The antibodies identified had specificity directed against antigens D, C, c and E in the Rh blood group system. The remainder had specificity directed against antigens Fy^a, K, Jk^a, M and Le^a. All antibodies identified by SPH-IAT, except 1 anti-M and 2 anti-Le^a, were also identified by manual methods. Blood donors: A positive reaction in the SPH-IAT test was noted in 0.7% of the samples. The identified antibodies had specificity directed against antigens D, C, E, Fy^a, K and M. All antibodies identified by the SPH-IAT, except 1 anti-D found in a sample from an RhD(+) male, were also identified by the manual methods. This study indicates that the SPH-IAT test is sensitive for detecting anti-M. This was also indicated for the SPH-ENZ test concerning the Jk^a antibody. An advantage of using solid-phase antibody screening tests for a larger series of samples is the possibility of automated sample identification, dilution, dispensing and interpretation of results. The SPH-IAT and the SPH-ENZ tests permitted us to select test cells, preferably from our own donor pool. We conclude that the SPH-IAT and SPH-ENZ tests are suitable for routine antibody screening of pregnant women and the SPH-IAT test for screening patients and blood donors.     

Reactivity of antinuclear antibodies with histones and other antigens in juvenile rheumatoid arthritis

Antinuclear antibodies are found in serum samples from most children with juvenile rheumatoid arthritis (JRA), but the antigenic specificities of these antibodies are not known. Using an immunoblot technique, we found that JRA patients' sera react with a variety of proteins in the nuclei of HEp-2 cells. Antibody to histone H1 was found in 42% of the JRA serum samples. An IgG antibody to a 45-kd protein was found in serum samples from some patients without uveitis, but it was not found in any sample from patients with uveitis. The immunoblot reactivity patterns do not appear to be useful in distinguishing between disease onset types or disease course types in patients with JRA.     

Comparative measurement of anti-factor VIII antibody by Bethesda assay and ELISA reveals restricted isotype profile and epitope specificity

Factor VIII (FVIII) inhibitor antibodies are produced against functional epitopes of FVIII in about 30% of severe hemophilia A patients leading to inhibition of its procoagulant activity. The Bethesda assay, the most commonly used method to measure FVIII inhibitors, based on inhibition of coagulant activity of FVIII, is neither able to detect noninhibitory antibodies nor their isotype. In this study we employed an indirect enzyme-linked immunosorbent assay (ELISA) to measure dif ferent isotypes and IgG subclasses of anti-FVIII anti body in the plasma of hemophiliacs (with and without inhibitor) and normal individuals using recombinant FVIII-coated microtiter plates. The results showed a predominance of IgG and IgG4, though IgA was slightly elevated in a few inhibitor-positive patients and IgM was hardly detectable. A highly significant correlation was found between the Bethesda titer and the optical density values of total Ig, IgG and IgG4 anti-FVIII antibodies obtained by ELISA (p<0.0001). These findings suggest a restricted specificity of anti-FVIII response in hemophiliacs towards functional epitopes of the molecule. Furthermore, high specificity and reasonable sensitivity of the ELISA, together with other technical advantages, suggest this method as a suitable supplementary technique for rapid large-scale screening of inhibitor-positive samples, though ELISA-negative samples need to be rechecked by the Bethesda assay to identify patients with a low inhibitor titer.     

不规则抗体筛选在新生儿溶血病中的临床诊疗

目的:探讨不规则抗体筛选在新生儿溶血病中的临床诊疗意义。方法:随机选取662例新生儿溶血病患儿,采用微柱凝胶免疫检测技术对其血清进行不规则抗体筛选。结果:共计检测有6例不规则抗体阳性结果。特异性抗体分别为抗-E 3例、抗-c 1例、抗-Ce 1例、抗-D 1例,6例患儿均为重症新生儿溶血病,经选用与患儿配合型血液换血治疗结合光疗和药物治疗,2周后痊愈出院。结论:不规则抗体筛选在新生儿溶血病诊断中意义重大,在协助诊断新生儿免疫性溶血性疾病方面优点尤其突出。早期适时对不规则抗体进行筛选,可以使重症新生儿溶血病患儿更早找到合适血源,减少输血等待时间,对保证新生儿输血安全十分重要。     

Eighteen Months'' Experience with a Manual Polybrene Crossmatch in a Large Hospital Transfusion Laboratory

For 18 months in this laboratory the manual polybrene technique (MP) has been used as the only crossmatching procedure preceded or accompanied by an antibody screen comprising two-stage papain and LISS antiglobulin techniques. There were 17,161 requests representing 43,006 blood units crossmatched and 20,841 units transfused. Non-specific reactivity with the MP required use of an antiglobulin crossmatch in approximately 0.2% of patient samples. Heparin in excess of 20 IU/ml reduced polybrene aggregation of red cells necessitating an antiglobulin crossmatch for 2 patients. Of 288 antibodies detected 20 reacted exclusively by MP compared with 18 by the papain procedure. The data supported the use of MP as an alternative to enzymes in antibody screening protocols. The polybrene technique was found to be a superior abbreviated crossmatch compared with the immediate spin technique and was applicable to all patients including those with known antibodies.     

Total plasma cholesterol in obesity after jejunoileal bypass with 3:1 or 1:3 jejunoileal ratio. A randomized trial.

Total plasma cholesterol (TPC) was measured repeatedly in 95 morbidly obese patients who had been randomized to non-operative management or jejunoileostomy with either a 3:1 or a 1:3 jejunoileal ratio. Initially, TPC was on average 6.4 mmol/l. and it remained stable in those not operated on. Within 1 month of surgery TPC decreased to a mean of 3.9 mmol/l, at which level it remained for the following 3 years. There was no difference in TPC between patients who had a 3:1 and those who had a 1:3 jejunoileal ratio of the functioning segment. Previous studies indicated that the increase in degradation of cholesterol to bile acids is much less in 1:3 than in 3:1 bypass. This study suggests that the changes in cholesterol metabolism after jejunoileostomy are dependent on the length of functioning jejunum and ileum in such a way that the effects of the two segments counterbalance each other.