Immunization, Isolation of Immunoglobulins and Antibody Titre Determination

Harboe, Niels; Ingild, A.

doi:10.1111/j.1365-3083.1983.tb04037.x

Cited by 150 publications

(74 citation statements)

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“…Thus, it was suggested that another type of RNase, showing no immunological cross-reactivity with anti-(RNase HS) antibody, which may be a secretory RNase, is present at low levels Activity is expressed as a pcrccntage of activity with no addition. The concentration of antibody was determined on the basis of the mass of the lyophilized IgC fraction purified by the method of Harboe and Ingild [24] in human spleen and scarcely at all in human liver. These findings are similar to the results described previously [3, 261.…”

Section: Immunological Propertiesmentioning

confidence: 99%

Purification and characterization of a ribonuclease from human spleen

Yasuda

Mizuta

Sato

et al. 1990

European Journal of Biochemistry

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A ribonuclease has been isolated from human spleen (RNase HS) by means of acid extraction, ammonium sulphate fractionation, successive column chromatographies on CM-cellulose, heparin-actigel, and poly(G)-agarose, and double gel-filtration on Sephadex G-75. The purified preparation was homogeneous as judged by SDSIPAGE. RNase HS was found to be a glycoprotein, containing three fucose, one mannose and five glucosamine residues/molecule, with a molecular mass of 17 kDa as determined by both SDS/PAGE and gel filtration. The catalytic properties and structural features, including its amino acid composition and the amino acid sequence of the N-terminal 35 residues, indicated that the enzyme was strictly related to nonsecretory RNase isolated from human urine and liver. In particular, the amino acid sequence of the N-terminal was identical with that of urine nonsecretory RNase and eosinophil-derived neurotoxin. Furthermore, analyses using three different antibodies specific to RNase HS, urine nonsecretory RNase and urine secretory RNase, indicated that RNase HS was not immunologically distinguishable from urine nonsecretory RNase, but clearly so from urine secretory RNase. However, the carbohydrate compositions of RNase HS and urine nonsecretory RNase were found to differ. It therefore remains to be resolved whether or not the tissue of origin of nonsecretory RNase in urine is the spleen.Sierakowska and Shugar [ 11 classified the pyrimidinespecific neutral and alkaline ribonuclease (RNase) of mammalian origin into two distinct classes : secretory and nonsecretory RNases. The former enzyme shows a pH optimum of around 8.5 and a greater preference for poly(C) than poly(U) and RNA. The latter cleaves RNA most effectively at pH 7.0, and its preference for poly(C) is less than the former.

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Section: Immunological Propertiesmentioning

confidence: 99%

Purification and characterization of a ribonuclease from human spleen

Yasuda

Mizuta

Sato

et al. 1990

European Journal of Biochemistry

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“…Moreover, the -production of polyclonal an-tibodies is difficult to standardize because it depends on the antigen used for immunization, the methods used, and the animal selected, its housing, and its nutrition. 10,14 Therefore, sufficient production, maximal standardization, and widespread applicability are better ensured by the use of MAbs as primary reagents in immunohistochemical techniques.…”

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confidence: 99%

Development of Murine Monoclonal Antibodies for the Immunohistochemical Diagnosis of Systemic Bovine Aspergillosis

Jensen¹,

Aalbæk

Lind³

et al. 1996

J VET Diagn Invest

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Abstract. Murine monoclonal antibodies (MAbs) against water-soluble somatic antigens (WSSA) and the wall fraction (WF) from Aspergillus fumigatus were produced by fusion of splenocytes from immunized BALB/c mice with mouse myeloma X63-Ag 8.653 cells. The supernatants of in vitro cultured hybridomas were initially screened for reactivity with the WSSA and the WF from A. fumigatus and WSSA of other fungi in an enzymelinked immunosorbent assay (ELISA). Supernatants reacting only with A. fumigatus antigens were subsequently screened for homologous and heterologous reactivity with immunohistochemical techniques using formalinfixed, paraffin-embedded tissues from experimentally infected mice. Because of a high immunohistochemical reactivity with homologous fungi, 4 MAbs raised against A. fumigatus WSSA and WF were selected for a further evaluation of cross-reactivity (diagnostic specificity) in immunohistochemical and immunoblotting assays. In immunohistochemical assays, all MAbs raised against WSSA cross-reacted heavily with a number of other fungal species. All 4 MAbs (MAb-WF-AF-1-4) raised against the WF reacted strongly with hyphae of Aspergillus spp.; hyphae of Scedosporium apiospermum were also strongly labeled by MAb-WF-AF-3 and-4. The 2 specifically reacting MAbs (MAb-WF-AF-1 and-2) were of the IgM biotype and were precipitating, and in immunoblotting experiments both bound to a 106-kD antigen of the WF, whereas they did not bind to WSSA of A. fumigatus. One of the 2 aspergillosis-specific MAbs (MAb-WF-AF-1) was used to screen 145 mycotic lesions of cattle. The diagnoses on bovine lesions obtained by MAb-WF-AF-1 were compared with results based on reactivity with heterologously absorbed polyclonal antibodies and, for some lesions, to culture results. In the vast majority of lesions (n = 133), the MAb-WF-AF-1 and the polyclonal anti-Aspergillus antibodies reacted in a similar pattern, i.e., positively in 41 aspergillosis lesions and negatively in 92 zygomycotic lesions. Hyphae in 3 of 12 lesions that were not stained by the polyclonal antibodies reacted with the specific MAb-WF-AF-1; i.e., aspergillosis was diagnosed. The characteristics of the 2 MAbs (MAb-WF-AF-1 and-2) raised against the WF of A. fumigatus in ELISA and immunoblotting and immunohistochemical assays justify their application for the in situ diagnosis of systemic aspergillosis of cattle.Aspergillus fuimigatus is the main cause of systemic bovine aspergillosis, a disease occuring worldwide and of great importance in cattle production. [2][3][4]14,[18][19][20][21]27,31,37 Because systemic mycoses including aspergillosis are difficult to diagnose clinically, most cases are not diagnosed until histopathologic evaluation is completed as part of a postmortem examination. 2-4,14,18-2l,27,3l,37 Attempts to culture the etiologic agents of systemic mycoses including Aspergillus spp. may give false results because of several difficulties, e.g., growth failure, overgrowth by contaminants, recovery of a fungus different from that suggested by the morpholo...

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“…Three months old female rabbits (Danish White, State Serum Institute, Denmark) were injected subcutaneously with 500 gg of osteocalcin absorbed to polyvinylpyrrolidone, PVP-40, (50% w/v in physiological saline) (17) emulsified with Freund's complete adjuvant. Three booster injections in Freund's incomplete adjuvant were given at 2-week intervals followed by injections at 4-week intervals (18). The titre was tested 10 days after the third boost by the ELISA described above.…”

Section: Biochemistry and Molecular Biology Internationalmentioning

confidence: 99%

Production and characterisation of monoclonal and polyclonal amtobpdoes agaomst bovine osteocalcin

et al. 1997

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SUMMARY: Monoclonal and polycl0nal antibodies were produced against osteocalcin purified from bovine bone. The antibodies were used for the development of a two-site ELISA. A monoclonal antibody recognising residues 15-49 of bovine osteocatcin generated by cleavage with 70% formic acid was used as capture antibody and a biotinylated polyclonal antibody recognising the same peptide was used for detection. The two-site assay did not detect any tryptic peptides of bovine osteocalcin (residues 1-19, 20-43, and 45-49) nor did it detect the peptide 1-14 or the synthetic peptide 37-49. Cross-reaction was observed for osteocalcin in plasma from sheep, goat, chicken and human but not for osteocalcin from rat and pig.

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Immunization, Isolation of Immunoglobulins and Antibody Titre Determination

Cited by 150 publications

References 0 publications

Purification and characterization of a ribonuclease from human spleen

Purification and characterization of a ribonuclease from human spleen

Development of Murine Monoclonal Antibodies for the Immunohistochemical Diagnosis of Systemic Bovine Aspergillosis

Production and characterisation of monoclonal and polyclonal amtobpdoes agaomst bovine osteocalcin

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