Fluorescent Antibody Method of Detecting Gonorrhea in Asymptomatic Females

Harris, Ad; Deacon, Wilbur E.; Tiedemann, John H.; Peacock, William L.

doi:10.2307/4591075

Cited by 13 publications

(6 citation statements)

References 1 publication

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“…technique was superior to conventional cultural diagnostic methods and afforded a considerable saving in labour and materials and in the time required for making a bacteriological diagnosis (Deacon, Peacock, Freeman, Harris, and Bunch, 1960). Similar studies on material from patients have been reported by Harris, Deacon, Tiedemann, and Peacock (1961), Lind (1962), Brown, Copeloff, and Peacock (1962), Moore, VanderStoep, Wende, and Knox (1963), and Danielsson (1963).…”

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confidence: 80%

Immunofluorescence Techniques as an Aid to the Diagnosis of Gonorrhoea

Fry¹,

Wilkinson²

1964

Sexually Transmitted Infections

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The growth from 18-hr cultures of four freshly-isolated strains of gonococci on chocolate agar slopes was pooled, washed twice with sterile saline, and resuspended in 4 ml. saline. This suspension was emulsified in an equal volume of Freund's complete adjuvant and 1 25 ml. of the emulsion was inoculated subcutaneously into the hind legs of young rabbits. A similarly prepared booster dose prepared from four further freshly-isolated strains of gonococci was given after 4 weeks and the animals bled out 3 weeks later after a trial had shown that complementfixing antibodies to a suspension of gonococci were present to a high titre.The globulin fraction of the antiserum was separated by half saturation with ammonium sulphate and after dialysis was conjugated with fluoresceine isothiocyanate by the method of Chadwick and Fothergill (1962), using a fluoresceine: protein ratio of 0 035 :1 0. Unreacted fluoresceine was removed from the conjugate by passage * Received for publication July 5, 1963. through a Sephadex G 25 column and elution with phosphate buffered saline, pH 7 3, followed by absorption with acetone-dried guinea-pig kidney tissue powder.On testing the conjugate against urethral smears from males with acute gonorrhoea, it was found that, while good labelling of gonococci was achieved, differentiation of the organisms from the background fluorescence of the leucocytes in the smears was difficult. Lind (1962) mentions the same problem. This was largely overcome by using normal rabbit globulin conjugated with Lissamine Rhodamine B as a background counterstain, as suggested by Smith, Marshall, and Eveland (1959). This conjugate was prepared by the method of Chadwick and Fothergill (1962) and for use was mixed with an equal volume of the fluoresceine-conjugated anti-gonococcal globulin. The proportions in which the two conjugates were mixed was determined for each batch made. The mixture gave a reddish-orange fluorescence to the leucocytes and other background material and afforded a good contrast to the green fluorescence of the labelled gonococci. Direct F.A. Test Thin smears of secretions, diluted if need be in tap water, were fixed by gentle heat and incubated with the mixed conjugates for one hour at 35 'C. in a moist chamber. After washing in two changes of buffered saline, pH 7 3, the slides were mounted in buffered glycerine.Delayed F.A. Test Secretions were inoculated on chocolate agar slopes in bijou bottles and incubated at 35'C. for 18 to 24 hrs. Smears of the mixed growth were fixed by heat and treated as in the direct test.Optical Equipment Slides were examined under dark-ground illumination using a Zeiss 1/ 12' objective with an iris diaphragm and a x 4 eyepiece. The illuminating source was a ME/D 250 watt high pressure mercury vapour lamp, light from which was passed through a 1 cm. trough of 5 per cent. CuSO4 to remove residual red light and a ChanceWatson OB 10 filter transmitting in the 340-550 m,u range. A Watson OY 12 barrier filter was used on the eyepiece.

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supporting

confidence: 80%

Immunofluorescence Techniques as an Aid to the Diagnosis of Gonorrhoea

Fry¹,

Wilkinson²

1964

Sexually Transmitted Infections

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“…In a num ber of women this infection m ust have taken an asym ptom atic course. This is not an unusual finding (Harris et al, 1961).…”

Section: Resultsmentioning

confidence: 83%

Diagnosis of Gonorrhea by a Fluorescent Antibody Technique

Mouton¹

1966

Dermatology

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“…'HE direct-staining fluorescent antibody (FA) technique has proved useful as a laboratory aid in the identification of Neisseria gonorrhoeae, or gonococcus (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)). An effective method for producing satisfactory fluorescein conjugated antiserums to this gonococcus (GC) has been found.…”

mentioning

confidence: 99%