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2-micron latex particles accumulated in macrophages in intestinal Peyer's patches of mice given latex suspensions as drinking fluid for 2 months. The number of particles accumulating was a direct (but nonlinear) function of the number ingested. Some of the latex particles were still present in Peyer's patches 6 weeks after the cessation of latex feeding.
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A method of preparation of papain (EC 3.4.22.2) from relatively soluble types of latex of Carica papaya, including spray-dried latex produced by a controlled and relatively mild process, was devised. Spray-dried latex dissolves easily in water up to 350mg/ml at 22 degrees C, which corresponds to approx. 230mg of protein/ml. When the usual method of preparation of crystalline papain contaminated only by its oxidation products, developed by Kimmel & Smith [J. Biol. Chem. (1954) 207, 515-531], is applied to spray-dried latex, the result is a preparation of papain heavily contaminated by chymopapains A and B (EC 3.4.22.6), and to a lesser extent by papaya peptidase A. This applies also to other types of papaya-latex currently commercially available, which, though less soluble than spray-dried latex, are more soluble than the types of latex available when the method of Kimmel & Smith (1954) was developed. This contamination is avoided by adjusting the concentration of the initial latex extract to 65mg of protein/ml (or less) before salt fractionation. For spray-dried latex this corresponds to 100mg of latex/ml. Papain isolated from spray-dried latex was characterized by using 2,2'-dipyridyl disulphide and 4-chloro-7-nitrobenzofurazan as thiol-specific reactivity probes and alpha-N-benzoyl-l-arginine ethyl ester as substrate. Papain isolated from this source appears to have the same catalytic-centre characteristics as papain isolated previously from latex produced by harsher methods. The catalysis of the hydrolysis of alpha-N-benzoyl-l-arginine ethyl ester by the mixture of thiol proteinases extracted from spray-dried latex by application of the method of Kimmel & Smith (1954) appears to obey Michaelis-Menten kinetics. The presence of the other enzymes results in an increase in the value of K(m) and a decrease in the catalytic-centre activity (k(cat.)) relative to the values for the catalysis by papain.
Natural rubber is harvested by periodically incising the laticifer vessels in the bark of rubber trees to release latex, the cytoplasm of laticifers. Although mitochondria are suspended in the cytoplasm and were expected to leak out, some earlier studies failed to detect their presence. In this study, we identified mitochondria and plastids in expelled latex using molecular methods and confocal microscopy, quantifying their abundance at 13 850 ± 800 mitochondria per microliter latex. Each tapping event released approximately 746 ± 35 mtDNA copies and 113 ± 7 mitochondria per laticifer cell (mean cell volume: 0.008134 μl). Individual mitochondria contained 6.7 ± 0.6 genome copies (mean ± SD), a value significantly higher than the 1.5 ± 0.2 mtDNA copies per mitochondrion observed in leaves. This suggests that laticifer mitochondria are primed for proliferation. We further investigated mitochondrial dynamics during tapping cycles by measuring temporal changes in concentration. Initial latex flow exhibited the highest mitochondrial concentration (18 749 ± 954/μl), which progressively decreased to 40% of the initial level (7542 ± 940/μl) within 30 min, likely due to dilution from water influx. Following latex vessel plugging, the mitochondrial population rebounded rapidly, surpassing the initial concentration by 1.4-fold within 3 days. Subsequent tapping cycles (second and third) exhibited similar mitochondrial loss and recovery trends, though recovery kinetics shifted from a linear (first cycle) to a logarithmic pattern. These results indicate that tapping stimulates mitochondrial proliferation and that laticifer mitochondria lack protective mechanisms comparable to those of the nucleus, resulting in their expulsion with latex during harvesting.
A commercially available test for systemic lupus erythematosus employing nucleoprotein-coated latex particles has been evaluated both clinically and serologically. The sera from all 40 subjects with active SLE had positive latex tests, while all 28 sera from healthy adults had negative latex tests. False-positive latex tests were observed in five of 13 antinuclear factor-positive patients with other chronic inflammatory diseases. Specific DNAase and RNAase digestion of the latex particles suggested the false-positive results were due to DNA moieties, other than native DNA, coating the latex particles.
Most samples of cerebrospinal fluid from 49 patients with or without coccidioidomycosis yielded positive agglutination with the Hyland ("CM") latex-particle agglutination test. Furthermore, simple dilution of sera from patients with or without coccidioidomycosis yielded a strong agglutination reaction with the latex particle agglutination test reagent. These reactions were found with three lots of the latex-particle agglutination test reagent. One lot latex particles was found to give agglutination reactions merely by addition of saline solution or water, though a second lot did not. It is advisable, therefore, that the latex-particle agglutination test not be applied to cerebrospinal fluid, and that it be applied cautiously, particularly with diluted sera.
Incubation of human platelets in plasma containing a suspension of latex particles for 1-90 min resulted in progressive accumulation of particles in the open-channel system, followed by localization of latex in electron-opaque vacuoles. After 60 min, acid phosphatase was localized within latex-containing vacuoles. The periodate-alkaline-bismuth reaction intensely stained external membranes and membranes of the open-channel system. Membranes of latex-containing organelles were not stained. Latex phagocytosis was independent of both anticoagulant choice and aspirin effects. Our results indicate that the platelet can act as a true phagocyte, and we suggest that the phagocytic process is chronologically similar to that reported for polymorphonuclear leukocytes.
The Polish synthetic latex LBS 3041 produced in Chemical Works "Oświecim" was examined for anatomical purposes. The latex was stained with pigments "Vulkan", pasts of general use "Pastokol" and commercial pigments to emulsive dyes. There was established a much better penetration into small blood vessels by latex LBS 3041 than by latex LBS 3060. On the account of the resistance of the examined latex to the coagulating activity of acetic acid, the hardening of the stuff after the injection of the specimens occurs through the cooling of preparations in the refrigerator.
The latex agglutination method, utilizing antibody-coated latex particles, was adapted for serogrouping of Neisseria meningitidis and serotyping of encapsulated Haemophilus influenzae strains from agar plates. It was found to give more clear-cut results than conventional slide agglutination. A 100% agreement with the antiserum agar method was found for all strains isolated from blood or cerebrospinal fluid. Many meningococcal strains from nasopharyngeal carriers are autoagglutinable, but some of these gave a positive reaction with the group B latex reagent, although they were negative by the antiserum agar method. The latex agglutination method has several advantages over others: the lack of autoagglutination, easy performance, easy interpretation, and very low consumption of antisera.
A new latex particle agglutination test (FIBROTEX) has been compared with the Thrombo-Wellcotest and the tanned red cell hemagglutination inhibition immunoassay (TRCHII) for sensitivity to purified fibrinogen and plasmic degradation products and for reactivity with 31 normal sera and 170 sera from patients with thrombotic disorders. The latex particles in the FIBROTEX are coated with antibodies against fibrinogen and fragment E (anti-FE), in contrast with the anti-fragment D and anti-fragment E antibodies which coat the latex particles of the Thrombo-Wellcotest (anti-DE). The relative sensitivities of the TRCHII and the anti-FE and anti-DE latex tests were, respectively, 0.5, 2.5 and 7.5 microgram/ml for fibrinogen; 0.5, 0.5 and 4.0 microgram/ml for fragment X; and 1.25, greater than 30 and 8.0 microgram/ml for fragment D. Compared with anti-DE, the anti-FE particles demonstrated fewer false-positive results with sera and urines of normals and higher titers, fewer false negatives and better overall correlation with the TRCHII results (r = 0.89 vs. 0.75) on pathologic sera. The FIBROTEX is simple and rapid (5 minutes) and provides a more sensitive and accurate determination of serum fibrin degradation products than the Thrombo-Wellcotest.
The principle of the latex agglutination technique was applied on the serodiagnosis of Chagas' disease. The latex particles coated with antigen from Trypanosoma cruzi are agglutinated by antibodies against T. cruzi in the serum of patients suffering thereof. In 11600 comparative determinations worked out mainly in a number of South American laboratories, the sensitivity of this test was compared with xenodiagnosis, CF, IFT and IHA. The latex Chagas test shows a mean coincidence of 90% with xenodiagnosis, 88% with CF, 75% with IFT, and 83% with IHA. The results indicate that the test is a useful new tool for the serodiagnosis of Chagas' disease. Also the data from CF, IFT and IHA were compared with each other. The latex Chagas test is sensitive even in the early stage of the disease. Also the specificity of the test was evaluated in other parasitic, bacterial and general diseases.
Studies have been carried out on the immunization of rats with antigen-coated latex particles. These studies confirmed that a significant lung blood eosinophilia is induced by a repeat intravenous injection of antigen-coated latex particles. Rats treated with antigen-coated latex particles also become sensitive to antigen such that on antigen challenge the animals show bronchoconstriction. Studies show that the bronchoconstriction is specific for antigen, but is unlikely to be mediated by tissue-fixing antibodies. Experiments comparing different immunization schedules involving latex particles indicate a strong association between the level of eosinophilia and degree of antigen-induced bronchoconstriction. These studies suggest that eosinophils may contribute to the anaphylactic bronchoconstriction.
Measles virus antigens covalently linked to latex spheres were used for measuring measles-specific antibodies in a direct agglutination test either in microtitre plates or as a rapid slide-agglutination test. The titres were compared to that obtained by conventional assays. The measles-latex spheres were also used as the antigen for a radio-immuno assay. By incorporating a 14C-radioactive marker into the measles-latex spheres their interaction with lymphocytes from multiple sclerosis and control patients was determined. Lymphocytes from multiple sclerosis patients reacted with a higher percentage of beads at low bead/lymphocyte ratios compared with controls, whereas the reverse was found when the ratio of beads was increased.
The photometric latex test (PLT) for the detection of rheumatoid factors (RF) has been correlated with the sensitized sheep cell test (SSC) and the latex slide test (LST). A total of 377 sera from patients with rheumatoid arthritis (RA) and 120 sera from control subjects were examined. When the PLT was carried out on the native sera at a lower buffer ionic strength (0.05 M), agglutination was noted in over 99% of cases, indicating interaction of complement with human immunoglobulin bound to the latex particle surface. Although thermal inactivation eliminated most of the complement agglutination, an increase in ionic strength (0.5 M) was found to be essential for measuring only RF agglutination. The serologic data were analysed statistically by computer. A highly significant correlation was found between all serologic tests and RA sera, but there was no such correlation with the control sera. The specificity of all tests was generally over 90% and did not vary significantly, but the sensitivity varied from 52.0% to 71.1%, confirming that about one-third of all RA patients are seronegative. The tests were analysed for their total diagnostic capability. The PLT with native sera at higher ionic strenght proved to be the most sensitive, but with thermally inactivated sera it had a better diagnostic capacity. The SSC test appeared to be less sensitive and of lower diagnostic validity. Rapid LST tests were clearly inferior to PLT tests. The PLT can be used as a reliable and straightforward serologic method of diagnosis in RA, especially when carried out at a higher ionic strength and with thermally inactivated sera. It should be given preference over other serologic tests for RF and could well become standard practice in rheumatologic serology as a substitute for the SSC test.
Fetuin bound latex spheres do not adhere to the membranes of non-infected cells but adhere to those of cells productively infected by fowl plague virus (FPV Dobson strain). In contrast, asialo fetuin spheres do not attach to the membranes of productively infected cells. Moreover latex fetuin spheres incubated with extracts of productively infected cells and extensively washed are specifically enriched in neuraminidase activity without any trace of haemagglutinin. These observations suggest that viral neuraminidase in the membrane is the site of attachment of the sialic acid moieties of fetuin spheres. These neuraminidase sites are detectable when L cells are productively infected by a mammalian cell adapted mutant of the Dobson strain (FPV-B) but are not detectable on L cells abortively infected by wild type (FPV+). However, even in the abortive system, neuraminidase is synthesised de novo as shown by its labelling with 14C-glucosamine and by its isolation from labelled extracts of infected cells by latex fetuin spheres. These results show that misintegration of viral neuraminidase in the plasma membrane of L cells is a feature of abortive infection of these cells by the Dobson strain of FPV. However the relationship (if any) of this misintegration to abortive infection remains to be established.
This report describes the sensitisation of latex particles and the use of this reagent on the Autotape system to select blood donations containing greater than or equal to 4 IU/ml of tetanus toxoid antibody. Plasma containing tetanus toxoid antibodies is processed to provide immunoglobulin for passive protection against tetanus. The techniques used for screening include immunoelectrophoresis [3], tetanus toxoid sensitised latex particles [5], haemagglutination [1] and automated haemagglutination [6]. The technique used by the Sheffield Regional Transfusion Centre until recently, was immunoelectrophoresis, selecting donors with 4 or more IU/ml. The Autotape system, previously described for brucellosis [4] and syphilis testing [2], provides a system for automated slide testing and a 5-min tetanus toxoid coated latex test has been developed to select suitable donations for the production of specific immunoglobulin.
Specimens of cerebrospinal fluid from 201 patients with meningitis caused by Neisseria meningitidis groups A/B/C/135, Streptococcus pneumoniae (23 types), and Haemophilus influenzae type b were tested for the presence of specific bacterial antigens by latex agglutination, counterimmunoelectrophoresis (CIE), and protein A co-agglutination. Specific antigens were found in 75% of the specimens by latex agglutination and CIE, and in 60% of the specimens by protein A co-agglutination. Non-specific reactions in protein A co-agglutination were prevented by heating the specimens to 100 degrees C for a few minutes. The three methods are simple and quick to perform. The smallest amount of antiserum was used in protein A co-agglutination, but we found this method less sensitive. Latex agglutination and CIE proved to be equally sensitive, but the first method was easier to adopt in practice.
Cultures of 3T3 cells, synchronised by serum deprivation, were shown to phagocytose latex beads of 0.81 micrometer diameter more avidly in the G1 phase of the cell cycle. The maximum rate of about 0.12 particles/cell/hour was recorded 12 h after stimulation with 10% fetal bovine serum. After 24 h, when mitosis was beginning, the uptake fell to just above the unstimulated level of 0.06 particles/cell/hour. Exocytosis was linear and only 7% of the total cell-associated latex was released in 30 h. Internalisation of latex beads was demonstrated by electron microscopy. No uptake was detected in experiments conducted at 4 degrees C.