scholarly journals Four unlabeled antibody bridge techniques: a comparison.

1981 ◽  
Vol 29 (12) ◽  
pp. 1397-1404 ◽  
Author(s):  
P Ordronneau ◽  
P B Lindström ◽  
P Petrusz
Keyword(s):  
Horseradish Peroxidase ◽  
Gamma Globulin ◽  
Staining Intensity ◽  
Reaction Products ◽  
Electron Microscopic ◽  
Microscopic Studies ◽  
Immunocytochemical Techniques ◽  
Unlabeled Antibody ◽  
Shape And Size

Four unlabeled antibody immunocytochemical techniques, the "single bridge" (Avrameas S: Immunocytochemistry 6:825, 1969; Mason TE, Phifer RF, Spicer SS, Swallow RS, Dreskin RD: J Histochem Cytochem 17:190, 1969a; Sternberger LA, Cuculis JJ: 1969), the "single peroxidase-antiperoxidase (PAP)" (Sternberger LA, Hardy PH Jr, Cuculis JJ, Meyer HG: J Histochem Cytochem 18:315, 1970), the "double PAP" (Vacca LL, Rosario SL, Zimmerman EA, Tomashefsky P, Ng P-Y, Hsu KC: J Histochem Cytochem 23:208, 1975) and the "double bridge" (Ordronneau P, Petrusz P: Am J Anat 158:491, 1980) were compared at both the light and electron microscopic levels. The "double" procedures involved repeating incubations with the bridge antibody, in this case, sheep anti-rabbit gamma globulin, followed either by a second PAP step for the "double PAP" or a second anti-horseradish peroxidase step and a single incubation in horseradish peroxidase for the "double bridge." At both the light and electron microscopic levels the staining intensity was greater with the "double" techniques than with the "single" ones. This is probably due to amplification achieved with the second sheep anti-rabbit gamma globulin step, permitting an increase in the number of horseradish peroxidase molecules bound for each molecule of tissue-bound primary antibody. Also, the quality of the various commercial PAP preparations tested was variable. With the weaker ones the staining intensity could be increased by performing an incubation in fresh horseradish peroxidase after the PAP step. Finally, in electron microscopic studies, the reaction products formed in both the bridge and PAP procedures were identical in shape and size.

scholarly journals ELECTRON MICROSCOPIC STUDY OF THE ADRENOCORTICOTROPIN-PRODUCING CELL WITH THE USE OF UNLABELED ANTIBODY AND THE SOLUBLE PEROXIDASE-ANTIPEROXIDASE COMPLEX

1972 ◽  
Vol 20 (8) ◽  
pp. 590-603 ◽  
Author(s):  
G. C. MORIARTY ◽  
N. S. HALMI
Keyword(s):  
Secretory Granules ◽  
Staining Intensity ◽  
Electron Microscopic ◽  
Early Effect ◽  
Acth Cell ◽  
Maximal Diameter ◽  
Soluble Peroxidase ◽  
Ultrathin Sections ◽  
Acth Secreting ◽  
Unlabeled Antibody

The technique involving use of unlabeled antibody and the peroxidase-antiperoxidase complex was used to identify the adrenocorticotropin (ACTH)-secreting cell in the anterior pituitary lobe of the rat and to localize ACTH in it electron microscopically in ultrathin sections. The ACTH cell is star-shaped, with processes extending around other cells, and contains secretory granules of a maximal diameter of 300 mµ arranged peripherally along the plasma membrane. Stain was observed on secretory granules, around them, in the Golgi complex and in rough endoplasmic reticulum. One day after adrenalectomy, the ACTH cell is degranulated and the staining intensity of its remaining granules and cytoplasm is decreased, suggesting release of ACTH stores. If cortisol is given 6 hr after adrenalectomy, 18 hr later the ACTH cells are well granulated and the granules stain more intensely than normal. In addition, staining around the granules and throughout the cytoplasm is more intense, suggesting that an early effect of cortisol is to block release of ACTH. Twenty-one days after adrenalectomy, the ACTH cells are greatly increased in numbers and have complex, tortuous processes filled with intensely stained secretory granules.

scholarly journals THE UNLABELED ANTIBODY ENZYME METHOD OF IMMUNOCYTOCHEMISTRY QUANTITATIVE COMPARISON OF SENSITIVITIES WITH AND WITHOUT PEROXIDASE-ANTIPEROXIDASE COMPLEX

1974 ◽  
Vol 22 (8) ◽  
pp. 782-801 ◽  
Author(s):  
JOHN P. PETRALI ◽  
DENNIS M. HINTON ◽  
GWEN C. MORIARTY ◽  
LUDWIG A. STERNBERGER
Keyword(s):  
Secretory Granules ◽  
Enzyme Reaction ◽  
Fold Increase ◽  
Immunocytochemical Staining ◽  
Intermediate Lobe ◽  
Reaction Products ◽  
Electron Microscopic ◽  
Specific Staining ◽  
Enzyme Method ◽  
Unlabeled Antibody

Araldite sections of rat pituitary intermediate lobe were used with anti-17-39 adrenocorticotropin in the unlabeled antibody enzyme method to compare electron microscopic immunocytochemical staining by peroxidase-antiperoxidase complex (PAP) with antiserum or purified antibody to peroxidase followed by peroxidase (PO). Quantitation of normalized optical densities of secretory granules offered high significance comparison (P < 0.0001) of experimental with control values and of experimental values with each other. Use of purified antibody (prepared by a new density gradient method) yielded four times higher sensitivity than antiserum to PO, while a 6.5-fold increase would have been expected from the degree of contamination of anti-PO in the serum by nonanti-PO immunoglobulin. Use of PAP was four to five times more sensitive than purified anti-PO and 20 times more sensitive than antiserum to PO. The increased sensitivity of PAP is explained by the high over-all binding affinity of PO for anti-PO in the cyclic PAP molecule, thus preventing the losses of PO that occur during washing when anti-PO and PO have been applied in sequence. Identification of the characteristic, cyclic PAP molecules affords confirmation of specific staining at high resolution. In the sequential application of anti-PO and PO, no PAP molecules are formed, thus making distinction of specific from nonspecific deposition of enzyme reaction products ambiguous. With the use of anti-17-39 ACTH and the intermediate lobe, the unlabeled antibody enzyme method was 16,000-100,000 times more sensitive than radioimmunoassay.

Cooling curve in production sweetened concentrated milk supplemented with whey: Influence on the size and microstructure of lactose crystals

2019 ◽  
Vol 25 (6) ◽  
pp. 451-461
Author(s):  
IT Smykov ◽  
AI Gnezdilova ◽  
YuV Vinogradova ◽  
AV Muzykantova ◽  
AK Lyamina
Keyword(s):  
High Speed ◽  
Cooling Curve ◽  
Electron Microscopic ◽  
Two Stage ◽  
Sweetened Condensed Milk ◽  
Cooling Regime ◽  
Condensed Milk ◽  
Microscopic Studies ◽  
Concentrated Milk

The aim of this work was to develop a methodology to calculate the cooling curve for the sweetened condensed milk with added whey powder production and to assess the cooling regime effect on the distribution of lactose crystals’ size and their microstructure. It is proposed to use a two-stage cooling curve. At the first stage, the cooling is carried out at a high speed, and at the second with a speed that varies depending on the rate of lactose crystallization. Electron microscopic studies have confirmed the cooling regime effect on the crystals’ microstructure. The practical use of the developed regime showed that the two-stage cooling regime allows to reduce the size of lactose crystals (P < 0.05) and improve the quality of the finished product.

Scanning electron microscopic studies on spermatozoa of anurans from India and Sri Lanka

2001 ◽  
Vol 22 (3) ◽  
pp. 303-308 ◽  
Author(s):  
Hareesh Joshy ◽  
Mitsuru Kuramoto
Keyword(s):  
Sri Lanka ◽  
Sperm Morphology ◽  
Sperm Head ◽  
Electron Microscopic ◽  
Frog Species ◽  
Scanning Electron Microscopic ◽  
Microscopic Studies ◽  
Generalized Type ◽  
Scanning Electron ◽  
Shape And Size

AbstractThe shape and size of spermatozoa of 11 frog species from India and Sri Lanka were examined by scanning electron microscopy. The spermatozoa of the genera Limnonectes and Euphlyctis were of the generalized type with a thick sperm head and a thin tail, whereas Indirana semipalmata had peculiar spermatozoa with a densely coiled sperm head and a thick tail. Rhacophorus microtympanum is likely to belong to the genus Philautus from sperm morphology. The spermatozoa of Microhyla ornata and Ramanella obscura were very similar, with a cone-shaped sperm head and a thin tail.

scholarly journals ULTRASTRUCTURAL IMMUNOCYTOCHEMISTRY WITH UNLABELED ANTIBODIES AND THE PEROXIDASE-ANTIPEROXIDASE COMPLEX A TECHNIQUE MORE SENSITIVE THAN RADIOIMMUNOASSAY

1973 ◽  
Vol 21 (9) ◽  
pp. 825-833 ◽  
Author(s):  
GWEN C. MORIARTY ◽  
C. MICHAEL MORIARTY ◽  
LUDWIG A. STERNBERGER
Keyword(s):  
Staining Intensity ◽  
Poor Quality ◽  
Immunocytochemical Staining ◽  
Great Promise ◽  
Electron Microscopic ◽  
Immunocytochemical Technique ◽  
Titration Curves ◽  
Rat Pituitary ◽  
Normal Rat ◽  
Unlabeled Antibody

Titration curves were developed with antisera to 17-39ACTH (adrenocorticotropin) and 1-39ACTH with the techniques of radioimmunoassay and electron microscopic immunocytochemistry. For the latter method, the unlabeled antibody peroxidase-antiperoxidase complex immunocytochemical technique was used to stain normal rat pituitary intermediate lobes. By radioimmunoassay standards, the 17-39ACTH antiserum was of poor quality. Its titration curve exhibited a flat slope and it did not bind a significant amount of labeled antigen beyond a 1: 30 dilution. However, immunocytochemical staining was detected with this antiserum at dilutions as high as 1:1,500. The antiserum to 1-39ACTH was of better quality by radioimmunoassay standards. It bound 45% of the labeled antigen at a dilution of 1:5,000. Immunocytochemical staining intensity was nearly maximal at a 1:5,000 dilution and decreased progressively to a limiting value at 1:16,000. However, when incubation times in the antisera were increased from 3 min to match those of the radioimmunoassay (48 hr) maximal staining was achieved at dilutions as great as 1:512,000 where only trace amounts of the labeled antigen were bound in the radioimmunoassay. It was concluded that the unlabeled antibody peroxidase-antiperoxidase complex immunocytochemical technique was sensitive enough to detect antibodies in sera of low titer and/or avidity which are not detected by a radioimmunoassay. The technique holds great promise for a sensitive assay system.

scholarly journals Conjugated avidin binds to mast cell granules.

1985 ◽  
Vol 33 (1) ◽  
pp. 27-32 ◽  
Author(s):  
M D Tharp ◽  
L L Seelig ◽  
R E Tigelaar ◽  
P R Bergstresser
Keyword(s):  
Mast Cell ◽  
Mast Cells ◽  
Horseradish Peroxidase ◽  
Staining Technique ◽  
Cellular Structures ◽  
Clinical Tool ◽  
Electron Microscopic ◽  
Simple Method ◽  
Human Mast Cells ◽  
Microscopic Studies

The glycoprotein, avidin, conjugated either to the enzyme horseradish peroxidase, or to the fluorochrome dyes, fluorescein or rhodamine, identifies the granules of mast cells in both tissues and cell suspensions. In the absence of prior fixation, mast cells were not identified with conjugated avidin; however, granules released from these cells were stained with this labeled glycoprotein. The specificity of avidin for mast cells was confirmed by the absence of conjugated avidin-positive cells in the skin of mice (S1/S1d) deficient in mature dermal mast cells. Electron microscopic studies confirmed that avidin binds specifically to individual mast cell granules rather than to other cellular structures. Rodent and human mast cells were readily stained with avidin conjugated to horseradish peroxidase or to either of the fluorochrome dyes. The conjugated avidin staining technique is a reliable and simple method for identifying rodent and human mast cells, one that is useful as both an investigative and a clinical tool.

Electron-microscopic studies of the CA antigen, epitectin

1986 ◽  
Vol 86 (1) ◽  
pp. 249-261
Author(s):  
M.E. Bramwell ◽  
G. Wiseman ◽  
D.M. Shotton
Keyword(s):  
Electron Microscopy ◽  
Monoclonal Antibodies ◽  
Electron Microscopic ◽  
Microscopic Studies ◽  
Terminal Site ◽  
Shape And Size

Epitectin, the mucin-like glycoprotein defined by the monoclonal antibodies CA1, CA2 and CA3, has been examined by electron microscopy to determine its shape and size. It appears to be a single extended strand with a mean length of about 270 nm. The antibodies CA1 and CA2 appear to bind preferentially to a terminal site on the epitectin molecule.

scholarly journals A modification of the unlabeled antibody enzyme method using heterologous antisera for the light microscopic and ultrastructural localization of insulin, glucagon and growth hormone.

1975 ◽  
Vol 23 (9) ◽  
pp. 666-677 ◽  
Author(s):  
S L Erlandsen ◽  
J A Parsons ◽  
J P Burke ◽  
J A Redick ◽  
D E Van Orden ◽  
...  
Keyword(s):  
Growth Hormone ◽  
Gamma Globulin ◽  
Cross Reactivity ◽  
Model Systems ◽  
Electron Microscopic Level ◽  
Immunocytochemical Staining ◽  
Electron Microscopic ◽  
Primary Antiserum ◽  
Enzyme Method ◽  
Unlabeled Antibody

The requirement of using homologous antisera (primary antiserum and peroxidase-antiperoxidase (PAP) complex raised in the same species) in the unlabeled antibody enzyme method has been investigated at the light and electron microscopic level using the localization of insulin, glucagon and growth hormone as model systems. Optimum immunocytochemical staining for all three antigens was observed when sheep or goat antirabbit gamma-globulin (S-ARgammaG or G-ARgammaG) were used to couple rabbit peroxidase-antiperoxidase complex with either guinea pig antisera to insulin (GP-AIS) or glucagon (GP-AGS), or monkey antisera to rat growth hormone (M-ARGH). The cross-reactivity between S-ARgammaG or G-ARgammaG and immunoglobulins in these primary antisera were substantiated by immunoelectrophoresis and radioimmunoassay. S-ARgammaG was shown to produce precipitation arcs with GP-AIS and M-ARGH that were similar to those seen when the latter were reacted with rabbit antiguinea pig gamma-globulin antiserum and goat antimonkey gamma-globulin antiserum, respectively. Radioimmunoassay results revealed that immunoprecipitation of 6-10% as compared to homologous antisera controls yielded excellent staining localization when S-ARgammaG was used for immunocytochemistry. Thus, heterologous antisera (primary antiserum and PAP complex raised in different species) may be used in the unlabeled antibody enzyme method as long as the coupling antiserum shows cross-reactivity with immunoglobulins of the primary antiserum and the PAP complex.

scholarly journals ELECTRON MICROSCOPIC STUDIES ON STREPTOCOCCI

1973 ◽  
Vol 138 (1) ◽  
pp. 245-258 ◽  
Author(s):  
John Swanson ◽  
Emil C. Gotschlich
Keyword(s):  
Cell Wall ◽  
Horseradish Peroxidase ◽  
Electron Microscopic ◽  
Streptococcal Cell Wall ◽  
Outermost Surface ◽  
Laminar Distribution ◽  
Specific Polysaccharide ◽  
Microscopic Studies ◽  
Group A ◽  
Protein Cell

The location of Group A carbohydrate in the streptococcal cell wall has been studied by several ultrastructural techniques. The findings, based largely on use of ferritin- and horseradish peroxidase-conjugated antibodies, are interpreted as demonstrating a discrete laminar distribution of the group-specific polysaccharide. This carbohydrate layer is located on the outermost surface of the cell wall in organisms lacking protein cell wall antigens.

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