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Allergens of the imported fire ant.

A whole body extact (WBE) was prepared by saline extraction of ground imported fire ants (IFA). A serum pool from rabbits immunized with the WBE elicited 31 precipitates on crossed immunoelectrophoresis. Crossed radioimmunoelectrophoresis was performed with sera from 26 RAST IFA-positive subjects, 24 of whom were known for clinical hypersensitivity to IFA and two subjects, known to be hypersensitive to yellow jacket. Radiostaining was obtained to 14 precipitates, some of which were judged to be partially identical. Thus, six apparently different allergen groups could be identified. Two of these fulfilled arbitrary criteria of major allergens. Three of the IFA WBE allergens were easily identified in crossed immunoelectrophoresis of a commercially obtained IFA-venom preparation. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the IFA WBE demonstrated protein bands with molecular weight ranging from 10,000 to 200,000 daltons. An immunoblotting technique with nitrocellulose paper strips revealed binding of human IgE from hypersensitive subjects to four different bands (either single bands or clusters of proteins of similar molecular weight) with molecular weights ranging from 13,000 to 42,000 daltons. It is a notable finding that these insect allergens thus were of a similar size as important aeroallergens.

Allergens↗

Inhibition of chronic relapsing experimental allergic encephalomyelitis in the Biozzi AB/H mouse.

Chronic relapsing experimental allergic encephalomyelitis (CREAE) can be reproducibly induced in Biozzi AB/H mice following injection of spinal cord homogenate (SCH) emulsified in complete Freund's adjuvant (CFA). Active clinical disease is associated with mononuclear cell infiltration of the central nervous system (CNS), mainly the spinal cord. Whole brain homogenate (BH), however, failed to induce clinical or histological disease. In contrast, substituting sciatic nerve homogenate in the inoculum induced experimental allergic neuritis (EAN). Clinical disease was manifest earlier (13.1 +/- 0.3 days) than CREAE (16.2 +/- 1.4) and was accompanied by mononuclear infiltration of the peripheral nervous system (PNS). In comparison to CREAE induction, pretreating mice with SCH or BH in incomplete Freund's adjuvant (IFA) suppressed the development of SCH-induced disease. The BH was more tolerogenic than the SCH and this hyporesponsiveness was CNS antigen-specific as PNS tissue failed to inhibit the course of CREAE. Tolerance induced by pretreatment with SCH or BH in IFA was reversed by a single injection of 200 mg/kg cyclophosphamide, 2 days prior to CREAE induction. This suggests that IFA-induced hyporesponsiveness is actively regulated, possibly via the action of suppressor cells. In addition, treatment with neuroantigens in IFA appears to be mainly afferent acting as it serves to prevent initial disease induction. This treatment after immunization for CREAE, however, fails to prevent disease progression. Furthermore, treatment with CNS antigens emulsified in IFA during the post-acute remission stage appeared to synchronize and induce (32 +/- 1 days) the onset of clinical relapse, compared with untreated controls (41 +/- 5 days). This indicates that such IFA treatment has minimal value in controlling an ongoing immune disease of the CNS.

Animals↗

Cryogenic preservation of virus-infected cells used as immunofluorescent assay substrates.

Cells infected with herpes simplex virus, respiratory syncytial virus and adenovirus were subjected to several cryogenic regimens to determine whether these procedures were detrimental to the performance and results obtained with an indirect immunofluorescence assay (IFA). Infected cells were harvested and divided into equal aliquots prior to freezing at different temperatures (-20, -70 and -165 degrees C). Cells fixed onto glass slides immediately after harvesting were used as controls. For the IFA, cells were thawed and fixed onto glass slides immediately prior to reaction with an appropriate virus-specific antibody and, subsequently, an FITC-conjugated anti-species antibody. The virus-infected cells retained their structural integrity following the experimental cryogenic regimens and were useable in the IFA. The quality of the image and the intensity of the fluorescent signal were enhanced by the incorporation of dimethyl sulfoxide into the freezing medium; cells frozen at -165 degrees C with DMSO gave the most satisfactory IFA results, although cells frozen at either -20 or -70 degrees C with DMSO were also useable. The use of cells frozen at the experimental temperatures in either phosphate buffered saline or tissue culture medium resulted in IFA substrate slides of diminished quality, often with a blurred image and distorted cellular morphology. We conclude that freezing of virus-infected cells prior to preparation for IFA is not detrimental to the performance of this procedure and the interpretation of immunological reactivity.

Adenoviridae↗

Influence of immunomodulatory agents on bovine humoral and cellular immune responses to parenteral inoculation with bovine rotavirus vaccines.

Sodium diethyldithiocarbamate (DTC), mycobacterium cell wall extract (MCWE, Regressin), killed Corynebacterium parvum (C. parvum, Immunoregulin) and muramyldipeptide (MDP) were each combined with purified, live bovine rotavirus and inoculated into 3 month-old Holstein-Friesian calves in order to examine their ability to potentiate specific humoral and cellular immune responses. The vaccinated calves were boosted twice at 3 and 6 weeks after initial vaccine inoculation. The rotavirus was administered intramuscularly either in an aqueous suspension or in a water-in-oil (WIO) emulsion, prepared with incomplete Freund's adjuvant (IFA). DTC and C. parvum were given by the intravenous route, while MCWE and MDP were incorporated directly in the rotavirus suspension. Two groups of calves were also vaccinated either with rotavirus and IFA or with rotavirus emulsified in mineral oil and a mannide oleate compound (MOC, Montanide 888). A control group of calves was given phosphate-buffered saline (PBS) solution emulsified with IFA. The different vaccine preparations were then compared by studying the kinetics of serum rotavirus-neutralizing antibody production and of proliferative response by blood lymphocytes following in vitro stimulation with bovine rotavirus. The results showed that: (1) the bovine rotavirus should be incorporated in a WIO emulsion in order to induce a cell-mediated immune response as detected by the rotavirus-specific in vitro stimulation test with blood lymphocytes, and to produce higher neutralizing antibody titers in the serum; (2) the vaccines prepared with the mineral oil-MOC complex or IFA both induced comparable levels of humoral and cellular immune responses. The use of mineral oil and MOC as adjuvant may be preferred to IFA, because of the facility of preparing the vaccine and of the low viscosity of the resulting WIO emulsion: (3) the addition of MDP to the WIO emulsion prepared with IFA resulted in a higher cell-mediated immune response as determined by the in vitro blood lymphocyte transformation index specific for bovine rotavirus.

Acetylmuramyl-Alanyl-Isoglutamine↗

Monoclonal antibodies to canine adenoviruses 1 and 2 that are type-specific by virus neutralization and immunofluorescence.

Monoclonal antibodies were produced against the Mirandola strain of canine adenovirus Type 1 (CAV-1) and the Manhattan strain of canine adenovirus Type 2 (CAV-2). The monoclonal antibodies were used in vitro in virus neutralization (VN) assays and in indirect fluorescent antibody (IFA) tests to examine several strains of each virus type. Out of 36 monoclonal antibodies produced against the Mirandola strain, 18 were type-specific for CAV-1 by IFA and 13 of those neutralized the virus in vitro. The other 18 antibodies bound both CAV-1 and CAV-2 by IFA; however, 7 of those specifically neutralized only CAV-1. The 160 monoclonal antibodies made against the Manhattan strain of CAV-2 yielded 77 type-specific antibodies by IFA, of which 39 neutralized only CAV-2 in vitro. The remaining 83 antibodies recognized both CAV-1 and CAV-2 by IFA, with 3 of those neutralizing both viral types. The hemagglutination inhibition (HI) test was performed on a selected monoclonal antibody from each specificity group. Although Type 1 CAV could be readily differentiated from Type 2 CAV by using type-specific monoclonal antibodies in the IFA or VN tests, strains within each type could not be differentiated. This is the first report of neutralizing monoclonal antibodies for a mammalian adenovirus.

Adenoviridae↗

Adaptation of a commercially available indirect fluorescent antibody slide test for measuring measles-specific immunoglobulins.

A commercially available indirect fluorescent antibody (IFA) test was evaluated for the determination of measles-specific immunoglobulins G and M (MIgG, MIgM). The IFA test detected fourfold rises of MIgG in 34 of 35 (97%) cases of measles confirmed by complement fixation or hemagglutination inhibition. In determining immune status, MIgG-IFA correlated with hemagglutination inhibition in 22 of 23 (96%) cases. The IFA test detected MIgM in only 11 of 34 acute-phase sera collected within 5 days of the reported onset of rash and in the convalescent specimens of another 13 of the 35 specimens. The IFA test is an effective method for the conventional diagnosis of measles and for determining immune status. This IFA test has a limited role as a rapid diagnostic test for measles when used to detect measles-specific MIgM in acute-phase sera obtained from patients with suspected measles.

Agglutination Tests↗

The clinical usefulness of a Puumalavirus recombinant nucleocapsid protein based enzyme-linked immunosorbent assay in the diagnosis of nephropathia epidemica as compared with an immunofluorescence assay.

BACKGROUND: Nephropathia epidemica (NE), a hemorrhagic fever with renal syndrome (HFRS) predominantly encountered in northern Europe, is a febrile disease, commonly associated with acute renal impairment. A rapid and reliable serological diagnosis is required to differentiate NE from other acute febrile illnesses in endemic areas. OBJECTIVE: To evaluate a Puumala (PUU) virus recombinant nucleocapsid protein (rN) based enzyme-linked immunosorbent assay (ELISA) for the serological diagnosis of NE as compared with an immunofluorescence assay (IFA) in a clinically relevant patient sample. STUDY DESIGN: During a four-month period, 618 serum samples from 512 patients with an illness suggestive of NE, sent to the Department of Clinical Virology for serological analysis, were included in the study. All sera were tested by PUU rN ELISA for presence of specific IgG, IgM and IgA antibodies and by IFA using PUU virus infected cells as antigen for presence of IgG and IgM antibodies. Patients with discordant results by IFA and rN ELISA were further serologically and/or clinically evaluated to assess the probability of NE. RESULTS: Compared to IFA, the specificities of the IgM and IgG rN ELISA were 100% and the corresponding sensitivities were 94.0%. The positive and negative predictive values of the PUU IgM rN ELISA in diagnosing NE infection was 100 and 98.6%, respectively. The positive predictive values for present NE infection of IgG rN ELISA and IFA were 68.3 and 71.4%, respectively. The positive predictive value of IgA rN ELISA was 95.8% and the negative 92.7%. CONCLUSIONS: The demonstration of specific IgM by rN ELISA is a highly specific and reliable method for the serological confirmation of NE. Detection of IgG antibodies by rN ELISA or IFA has a low predictive value to diagnose NE in an endemic area. The diagnostic value of IgA determination is in between IgM and IgG determinations.

Journal Article↗

Evaluation of an in-practice test for feline coronavirus antibodies.

A commercially available in-practice test for feline coronavirus (FCoV) antibodies (FCoV Immunocomb, Biogal Galed Laboratories) was evaluated by comparison with the gold standard FCoV immunofluorescent antibody (IFA) test. One hundred and three serum or plasma samples were selected and tested: 70 were positive by both tests, 24 were negative by both tests. The in-practice test produced five false positive and four false negative results. The sensitivity of the in-practice test was 95% and the specificity was 83%. When the titres were compared it was found that the in-practice test results were significantly correlated with IFA titres but the degree of correlation was not likely to be clinically useful. The IFA titres of the four false negative samples were found to be low (less than 40) which suggests that even a cat with a false negative result is still unlikely to be excreting FCoV. A negative result with the in-practice assay is likely to be reliable for screening cats prior to entry into an FCoV-free cattery or stud. It would also be useful in the investigation of suspected FIP as most cats with this condition have high IFA titres of antibodies. A strong positive result would be useful in the diagnosis of FIP (in conjunction with other biochemical and cytological testing), but positive results would be of limited value in monitoring FCoV infection in healthy cats as the antibody titre could not be reliably compared with those obtained with IFA. All positive results obtained using the in-practice kit should be confirmed and titrated by IFA. The kit also appeared to work efficiently with ascites samples (n=6) but too few samples were analysed to draw firm conclusions.

Animals↗

The natural history of exposure to the imported fire ant (Solenopsis invicta).

BACKGROUND: Imported fire ants (IFA) are a common cause of insect venom hypersensitivity in the southeastern United States. The purpose of this study was to determine the sting attack rate and development of specific IgE in an unsensitized population. METHODS: Study participants consisted of 137 medical students with limited exposure to IFA-endemic areas who were temporarily training in San Antonio, Tex. Subjects were surveyed for prior IFA exposure with a questionnaire, and IFA-specific IgE was evaluated with RAST and intradermal skin testing. Evaluations were performed on arrival and reported at departure from the endemic area 3 weeks later. RESULTS: One hundred seven subjects completed the study. Field stings were reported in 55 subjects, resulting in a sting attack rate of 51%. In these 55 subjects 53 (96%) reported a pustule or a small local reaction at the sting site, one (2%) reported an isolated large local reaction, and none reported a systemic reaction. At the 3-week follow-up skin test and RAST conversions occurred in seven subjects (13%) and in one subject (1.8%), respectively. CONCLUSIONS: Even brief exposures to IFA-endemic areas result in significant sting rates and concurrent rapid development of IFA-specific IgE in 16% of stung subjects.

Adult↗

Adjuvant activity of incomplete Freund's adjuvant.

The water-in-oil emulsion adjuvants developed by Jules Freund and coworkers have been, and continue to be, used in research and veterinary vaccines. Incomplete Freund's Adjuvant (IFA) has also been widely tested in humans where it was shown to augment antibody responses greater than other adjuvants, such as alum, and to be well tolerated. Toxicity associated with the use of IFA has been controlled by the use of high grade oils and purified surfactants and clinical testing is ongoing to day. One potential limitation of IFA is that it does not augment the types of cellular immune responses that thought to be critical to the control of many viral infections and tumors. However, IFA can be mixed with or used in conjunction with other adjuvant-active compounds, such as saponins and cytokines. These mixed formulations or vaccine protocols have proved successful in animal testing and may soon be tested in clinical trials. Thus, the toxicity and potential limited adjuvant activity of IFA can be controlled or corrected making IFA a useful product in the vaccine industry.

Journal Article↗

Seroepidemiologic studies on Babesia equi and Babesia caballi infections in Brazil.

Horses from six stud farms representing the most frequent types of horse breeding in Brazil were tested for Babesia antibodies by the IFA test. The farms are located at the tropic of Capricorn at an altitude of 472-715 m where temperatures below 0 degrees C may occur. Horses of conventional stud farms were infested with Dermacentor nitens, Amblyomma cajennense, and Boophilus microplus. Infestation with Bo. microplus was associated with direct or indirect contact of horses with cattle, and was not detected at professional stud farms. At one large professional stud farm, only D. nitens was observed. Prevalence of Babesia equi correlated positively (p<0.001) with contact of pastured horses to cattle (67.1% versus 17.5%). The IFA test was validated using sera from 92 non-infected horses and from 18 ponies infected experimentally with the USDA strains of B. equi or B. caballi or with the Brazilian isolates from the study area. Differences in test results obtained using antigens from USDA strains or Brazilian isolates were not significant. The specificity was 100% except for the IFA test using Brazilian antigen of B. caballi (98%). The sensitivity was 100% except for the IFA test using the USDA antigen of B. caballi. Accuracy of the tests ranged from 98 to 100%, and predictive values from 99 to 100%. Only 59% (132/224) of sera, positive by the IFA test for B. equi, tested positive by CF test, and 45% (232/515) of sera, positive by the IFA test for B. caballi, also tested positive by CF test. In all, 740 field sera from 140 horses, including 63 mares and their foals, were tested. Prevalence and incidences of B. caballi infections were significantly higher than those of B. equi infections: 49.2% (31/63) of the mares were infected with B. equi, but 79.4% (50/63) with B. caballi; 36% (18/50) of the foals became infected with B. equi within 12 months, but 100% (50/50) with B. caballi within 10 months. Maternal antibodies against B. equi and B. caballi in foals were 44 (22/50) and 68% (34/50), respectively. Titers persisted for 1-5 months for B. equi and 1-4 months for B. caballi.

Animals↗

Natural history of large local and generalized cutaneous reactions to imported fire ant stings in children.

BACKGROUND: There are no published data on the natural history of large local and generalized cutaneous reactions to imported fire ant (IFA) stings in children. OBJECTIVE: To determine the natural history of large local and generalized cutaneous reactions to IFA stings in children not treated with immunotherapy by reviewing medical records from a venom clinic during the past 20 years. METHODS: Patients were selected for the study if they were 16 years or younger at the time of the reaction, had only cutaneous symptoms (generalized cutaneous or large local), and did not initiate immunotherapy. Each patient's parents were asked to recall whether the patient had experienced any further stings since the last evaluation at the venom clinic. RESULTS: We contacted 31 of 57 patients evaluated between July 10, 1984, and February 5, 2004. Twenty patients (65%) reported that they had not developed more severe reactions with subsequent stings. Reactions remained cutaneous only. Eleven patients (35%) had not been stung again since the original evaluation. Five of these patients had moved out of the IFA-endemic region. None of the previously evaluated patients reported subsequent life-threatening anaphylaxis from IFA stings. CONCLUSIONS: These limited data on IFA stings suggest a benign outcome in children 16 years and younger with large local or generalized cutaneous reactions. Larger and more extensive studies need to be conducted to further define the natural history of cutaneous reactions to IFA stings in children.

Adolescent↗

Effect of imported fire ant extract on the degradation of mountain cedar pollen allergen.

BACKGROUND: Dust mite, cockroach, and mold extracts have been shown to contain proteases capable of degrading the proteins in other extracts. Loss of potency of allergens has been reported in mixtures containing cockroach and fungal extracts. Fire ant venoms consist of 90% to 95% n-alkyl and n-alkenyl piperidine alkaloids, which are not allergenic. No studies are available addressing the mixture of imported fire ant (IFA) whole-body extract with other allergens or the presence of proteolytic activity in the venom extract. OBJECTIVES: To evaluate the stability of mountain cedar pollen extract mixed with IFA whole-body extract and to qualitatively analyze the extract mixture for degradation of mountain cedar protein. METHODS: One milliliter each of mountain cedar and IFA whole-body extracts at a concentration of 500 microg/mL were combined and stored at 4 degrees C for 1, 3, 6, 15, 30, 60, 90, and 180 days. Separate mixtures of 1 mL of mountain cedar and IFA with 1 mL of human serum albumin were used as controls. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis was performed, and protein bands were qualitatively analyzed for degradation. RESULTS: We detected 3 distinct IFA protein bands and 1 mountain cedar protein band. With respect to these bands, no protein degradation was observed during 6 months of study in the extract mixture compared with the controls. CONCLUSIONS: Imported fire ant whole-body extract does not seem to degrade mountain cedar protein. Mixtures of allergenic extracts may be able to include IFA whole-body extract.

Allergens↗

Biological control of fire ants: an update on new techniques.

OBJECTIVE: To review the present understanding of biological control methods for imported fire ants (IFAs). DATA SOURCES: We searched MEDLINE, Biological Abstracts, and the US Department of Agriculture Formis Ant Literature database. STUDY SELECTION: All articles published in the last 10 years on biological control of fire ants were selected. RESULTS: The decapitating flies Pseudacteon tricuspis, Pseudacteon curvatus, and Pseudacteon litoralis have been successfully released in the United States. The continued releases of multiple species of decapitating flies will expand the area of impact, applying greater pressure on IFA populations throughout the southern United States. The microsporidium Thelohania solenopsae causes the slow demise of a fire ant colony. The advantages of T. solenopsae as a biological control agent include debilitation of queens, specificity for IFAs, self-sustaining infections, and lower relative tolerance to chemical pesticides. Solenopsis daguerrei has also been shown to have detrimental effects on IFA colony growth, the number of sexual reproductives produced, and the number of host queens in multiple queen colonies; however, this parasite is difficult to rear in the laboratory and to introduce into IFA colonies. CONCLUSIONS: It is unlikely that IFAs can be completely eradicated from the United States. However, technology using chemicals and/or natural control agents could eventually maintain populations at low levels if an integrated approach is used for control.

Animals↗

Evaluation of an indirect fluorescence immunoassay for the detection of serum antibodies against Babesia divergens in humans.

Since an indirect fluorescence immunoassay (IFA) for the detection of specific antibodies against Babesia divergens in human sera is not commercially available, an in-house prepared B. divergens IFA for the examination of bovine sera was established for serological studies in humans. To determine whether the described IFA is appropriate for such studies, 2 B. divergens antigens (of human or bovine origin) were tested using serum samples obtained from febrile human patients with a history of 'tick bite'. Sera from other species of animals infected with B. divergens, Babesia EU1, B. microti or B. canis were also included for comparative purposes. All serum samples were also tested using a commercially available IFA for the detection of antibodies against B. microti, and the results compared with those obtained using blood smear and molecular techniques. This study showed that the evaluation and standardization of a B. divergens IFA for testing human sera is critical and that different B. divergens antigens provide different end-point titres of antibodies, leading to false negative or positive results. Serological cross-reactivity between B. divergens and Babesia EU1 needs to be taken into account when interpreting IFA results.

Animals↗

Frequency of anti-bactericidal/permeability-increasing protein (BPI) and anti-azurocidin in patients with renal disease.

The major subtypes of anti-neutrophil cytoplasmic antibodies (ANCA) detected by indirect immunofluorescence assay (IFA) are P-ANCA and C-ANCA. In patients with vasculitis, myeloperoxidase (MPO) is the major P-ANCA antigen and proteinase 3 (PR3) is the major C-ANCA antigen. BPI and azurocidin, which are also called 57-kD cationic antimicrobial protein (CAP 57) and 37-kD cationic antimicrobial protein (CAP 37), respectively, have been proposed as less frequent target antigens for C-ANCA and P-ANCA. In patients with renal disease, we determined the frequency of antibodies against BPI and azurocidin. By IFA on alcohol-fixed neutrophils, monoclonal and polyclonal anti-BPI antibodies produced a C-ANCA pattern, whereas rabbit anti-azurocidin antibody produced a P-ANCA pattern. By ELISA, sera from 229 P-ANCA-positive patients, 99 C-ANCA-positive patients and 48 ANCA-negative (by IFA) patients with renal biopsies were tested for reactivity with recombinant human BPI and purified human azurocidin. Of these sera, 17.5% of P-ANCA, 30.3% of C-ANCA and 20.8% of IFA-ANCA-negative sera were positive for anti-BPI; and 8.3% of P-ANCA, 3.0% of C-ANCA and 8.3% of IFA-ANCA-negative sera were positive for anti-azurocidin. There was no statistical difference in frequency of anti-BPI between pauci-immune necrotizing and crescentic glomerulonephritis (NCGN) and other glomerular disease (OGD), and there was a lower frequency of anti-azurocidin in NCGN samples than in OGD samples. By Western blot, anti-BPI-positive sera reacted with a 57-kD BPI band and anti-azurocidin-positive sera with a 29-kD azurocidin band. In conclusion, there is a low frequency of anti-BPI and anti-azurocidin antibodies in ANCA-positive patient sera; however, this does not correlate with NCGN, which is a marker for ANCA-associated small vessel vasculitis, and a similar positivity is found in IFA-ANCA-negative patients with renal disease. Therefore, serologic detection of anti-BPI and anti-azurocidin is not diagnostically specific in patients with renal disease.

Amino Acid Sequence↗

An evaluation of four methods for the detection of Pneumocystis carinii in clinical specimens.

We undertook a prospective evaluation of 4 methods for the detection of Pneumocystis carinii in clinical specimens and compared an indirect immunofluorescence assay (IFA) (Diagnostics Pasteur), and a fluorescent whitening agent (FWA) (Blankophor BA 267%, Bayer, Australia) with our standard methenamine silver (MeAg) and toluidine blue O (TB) stains. Two hundred and two specimens were received from 162 patients (133 HIV infected, 19 heart or heart-lung transplant recipients, and 10 "miscellaneous"). The specimens consisted of 132 induced sputa, 56 bronchoalveolar lavage specimens, 10 fine needle aspiration lung biopsies, and 4 pleural fluid specimens. P. carinii was detected in 44 (22%) of the specimens. The sensitivities for the detection of P. carinii pneumonia were IFA: 92% (95% CI, 83-100%), FWA: 57% (95% CI, 41-73%), MeAg: 54% (95% CI, 38-70%), and TB: 49% (95% CI, 33-65%). Discordant results were greatest in specimens from patients who were receiving specific anti-P. carinii prophylaxis, or who had received treatment for several days prior to sampling. IFA was the most sensitive test and relatively easy to perform. IFA was also the most expensive test. We found the FWA method a useful screening test as it is cheap and quick to perform. However, it is less sensitive than IFA, which should be performed on the negative specimens. With the increasing use of specific anti-P. carinii prophylaxis in HIV-infected patients, methods more specific and sensitive than MeAg and TB stains are required. We have found IFA to improve significantly the rate of detection of P. carinii in this patient group.

Evaluation Studies as Topic↗

Stereoselectivity in metabolism of ifosfamide by CYP3A4 and CYP2B6.

The aim was to identify the hepatic cytochromes P450 (CYPs) responsible for the enantioselective metabolism of ifosfamide (IFA). The 4-hydroxylation, N2- and N3-dechloroethylation of IFA enantiomers were monitored simultaneously in the same metabolic systems using GC/MS and pseudoracemate techniques. In human and rat liver microsomes, (R)-IFA was preferentially metabolized via 4-hydroxylation, whereas its antipode was biotransformed in favour of N-dechloroethylation. CYP3A4 was the major enzyme responsible for metabolism of IFA enantiomers in human liver. The study also revealed that CYP3A (human CYP3A4/5 and rat CYP3A1/2) and CYP2B (human CYP2B6 and rat CYP2B1/2) enantioselectively mediated the 4-hydroxylation, N2- and N3-dechloroethylation of IFA. CYP3A preferentially supported the formation of (R)-4-hydroxyIFA (HOIF), (R)-N2-dechloroethylIFA (N2D) and (R)-N3-dechloroethylIFA (N3D), whereas CYP2B preferentially mediated the generation of (S)-HOIF, (S)-N2D and (S)-N3D. The enantioselective metabolism of IFA by CYP3A4 and CYP2B1 was confirmed in cDNA transfected V79 cells.

Animals↗