Effects of primary antiserum dilution on staining of "antigenrich" tissues with the peroxidase antiperoxidase technique.

1977 ◽  
Vol 25 (6) ◽  
pp. 443-447 ◽  
Author(s):  
J W Bigbee ◽  
J C Kosek ◽  
L F Eng
Keyword(s):  
Optic Nerve ◽  
Antigen Binding ◽  
Frozen Sections ◽  
Primary Antiserum ◽  
Rat Cerebellum ◽  
Antiserum Dilution ◽  
Formalin Fixed

The effect of primary antiserum dilution on staining results with the peroxidase antiperoxidase method were investigated using frozen sections of perfused rat cerebellum and optic nerve. Results comparable to formalin fixed and paraffin embedded tissue were attainable only when low antiserum concentrations were used. Optimal staining of antigen rich tissue, such as frozen sections, with the peroxidase antiperoxidase method required low antiserum concentrations apparently to minimize the binding of both antigen-binding fragments of the bridging antibody to the tissue bound antiserum. It appears that low antiserum concentration insures that sufficient bridge antibody molecules will be only singly bound and thus free to attach the peroxidase antiperoxidase complex.

Ultrastructural Localization of Acid Mucosubstance in Skeletal Muscle

1967 ◽  
Vol 25 ◽  
pp. 52-53 ◽  
Author(s):  
William J. Dougherty ◽  
Samuel S. Spicer
Keyword(s):  
Striated Muscle ◽  
Divalent Cations ◽  
Ultrastructural Localization ◽  
Major Elements ◽  
Frozen Sections ◽  
Thorium Dioxide ◽  
Iron Solution ◽  
Coupling System ◽  
Psoas Muscles ◽  
Formalin Fixed

In recent years, considerable attention has focused on the morphological nature of the excitation-contraction coupling system of striated muscle. Since the study of Porter and Palade, it has become evident that the sarcoplastic reticulum (SR) and transverse tubules constitute the major elements of this system. The problem still exists, however, of determining the mechamisms by which the signal to interdigitate is presented to the thick and thin myofilaments. This problem appears to center on the movement of Ca++ions between myofilaments and SR. Recently, Philpott and Goldstein reported acid mucosubstance associated with the SR of fish branchial muscle using the colloidal thorium dioxide technique, and suggested that this material may serve to bind or release divalent cations such as Ca++. In the present study, Hale's iron solution adapted to electron microscopy was applied to formalin-fixed myofibrils isolated from glycerol-extracted rabbit psoas muscles and to frozen sections of formalin-fixed rat psoas muscles.

scholarly journals A simple technique for preservation of fixation-sensitive antigens in paraffin-embedded tissues.

1994 ◽  
Vol 42 (8) ◽  
pp. 1127-1134 ◽  
Author(s):  
J H Beckstead
Keyword(s):  
Simple Technique ◽  
Frozen Section ◽  
Lymphoid Tissues ◽  
Frozen Sections ◽  
Morphological Examination ◽  
Fixed Tissue ◽  
Formalin Fixed Tissue ◽  
Important Approach ◽  
Reduced Toxicity ◽  
Formalin Fixed

Immunohistochemistry is a powerful tool for tissue diagnosis and research. Although the frozen section has remained the gold standard for this important approach to evaluation of antigens in tissues, there is widespread acknowledgment of many limitations. Routine paraffin-embedded sections ware widely used for morphological examination of tissues but are not optimal for antigen preservation. In this study, paraffin-embedded tissues fixed with a simple buffer containing zinc as the primary fixative were compared with tissues fixed with routine formalin, zinc-formalin, paraformaldehyde, ethanol, a variety of commercial (non-formalin-containing) fixatives that have been recommended for reduced toxicity and improved antigen survival, and frozen sections. Human lymphoid tissues and a group of antibodies to antigens (CD1, CD4, CD7, CD8, CD19) usually preserved only in frozen tissue were used as a model system. Fixation in a simple solution of zinc acetate and zinc chloride in a Tris-Ca acetate buffer resulted in antigen preservation comparable to that in frozen sections with antibodies to these cell surface markers. Morphological preservation was comparable to formalin-fixed sections. The work presents a new method that represents the closest approach yet to a technique that combines optimal antigenic survival with the convenience and morphological preservation of traditional formalin-fixed tissue embedded in paraffin.

METACHROMATIC LEUKO-ENCEPHALOPATHY

PEDIATRICS ◽  
1958 ◽  
Vol 22 (6) ◽  
pp. 1064-1073
Author(s):  
Raymond F. Hain ◽  
Gerald D. LaVeck
Keyword(s):  
Illustrative Case ◽  
Frozen Sections ◽  
Paraffin Sections ◽  
Mental Deterioration ◽  
Pathologic Findings ◽  
Blue Stain ◽  
The Central Nervous System ◽  
Metachromatic Granules ◽  
The Brain ◽  
Formalin Fixed

Metachromatic leuko-encephalopathy is a familial degenerative disease of the central nervous system included with Schilder's disease as a type of diffuse cerebral sclerosis. The disease usually has its onset early in childhood and is characteried by progressive motor and mental deterioration with ataxia, muscular weakness, spasticity, optic atrophy, convulsions and finally dementia. The concentration of protein of the cerebrospinal fluid is frequently elevated. The pathologic findings consist of demyelination, destruction of axons, gliosis and the accumulation of metachromatic granules in the brain and other organs. The metachromasia can be demonstrated in formalin-fixed frozen sections with a toluidine blue stain. It is not demonstrable in paraffin sections. Histochemical studies indicate this abnormal material is probably a complex of glycolipids and protein. It has been reported elsewhere that early diagnosis can be established by demonstrating the metachromatic materials in urinary sediment or in a real biopsy. An illustrative case has been presented.

Improvement of the Quality of Frozen Sections from Formalin fixed Tissue

1990 ◽  
Vol 65 (1) ◽  
pp. 43-44
Author(s):  
Toshihiro Ishii ◽  
Kumiko Kasama ◽  
Mayumi Kondo
Keyword(s):  
Frozen Sections ◽  
Fixed Tissue ◽  
Formalin Fixed Tissue ◽  
Formalin Fixed

The Histochemical Localization of Cholesterol in Formalin-Fixed and Fresh Frozen Sections

1976 ◽  
Vol 51 (3) ◽  
pp. 143-145 ◽  
Author(s):  
D. J. Rossouw ◽  
Carol C. Chase ◽  
Ina Raath ◽  
F. M. Engelbrecht
Keyword(s):  
Histochemical Localization ◽  
Frozen Sections ◽  
Fresh Frozen ◽  
Formalin Fixed

scholarly journals CYTOCHEMICAL OBSERVATIONS ON THE RELATIONSHIP BETWEEN LYSOSOMES AND PHAGOSOMES IN KIDNEY AND LIVER BY COMBINED STAINING FOR ACID PHOSPHATASE AND INTRAVENOUSLY INJECTED HORSERADISH PEROXIDASE

1964 ◽  
Vol 20 (3) ◽  
pp. 497-507 ◽  
Author(s):  
Werner Straus
Keyword(s):  
Acid Phosphatase ◽  
Egg White ◽  
Frozen Sections ◽  
Weak Reaction ◽  
Tubule Cells ◽  
Portal Veins ◽  
Color Reactions ◽  
Relationship Of ◽  
The Relationship ◽  
Formalin Fixed

After incubation of formalin-fixed, frozen sections of kidney and liver from peroxidase-treated rats in an azo dye medium for acid phosphatase, and after subsequent incubation of the same sections with benzidine, phagosomes were stained blue and lysosomes were stained red in the same cells. It was observed that newly formed phagosomes were separate from preexisting lysosomes in the tubule cells of the kidney and in the Kupffer cells of the liver at early periods after treatment with peroxidase. At later periods, the color reactions for acid phosphatase and peroxidase occurred in the same granules. The reaction of peroxidase decreased gradually and disappeared from the phago-lysosomes after 2 to 3 days, whereas the reaction for acid phosphatase persisted. In the liver, most of the injected protein was concentrated in large phagosomes located at the periphery of the cells lining the sinusoids. The peribiliary lysosomes showed a relatively weak reaction for peroxidase in the proximity of the portal veins. After pathological changes of permeability, phagosomes and lysosomes lost their normal location and fused, in the interior of many liver cells, to form large vacuoles or spheres. The effects of a reduced load of peroxidase and the effects of the pretreatment with another protein (egg white) on the phago-lysosomes of the kidney were tested. The relationship of the fusion of phagosomes with lysosomes to the size of normal and pathological phago-lysosomes was discussed.

scholarly journals Effect of Fixation and Epitope Retrieval on BrdU Indices in Mammary Carcinomas

2000 ◽  
Vol 48 (3) ◽  
pp. 355-362 ◽  
Author(s):  
John N. McGinley ◽  
Katrina K. Knott ◽  
Henry J. Thompson
Keyword(s):  
Cell Proliferation ◽  
Prospective Studies ◽  
Immunohistochemical Staining ◽  
Staining Technique ◽  
Labeling Index ◽  
Antigen Retrieval ◽  
Frozen Sections ◽  
Mammary Carcinomas ◽  
Incorporated Brdu ◽  
Formalin Fixed

Studies in which 5-bromo-2′-deoxyuridine (BrdU) is used to quantify rates of cell proliferation are conducted prospectively. Therefore, the opportunity exists to select conditions that optimize detection of the BrdU epitope. The objective of this study was to quantify the extent to which the BrdU epitope was masked by formalin vs methacarn fixation in the assessment of cell proliferation. Mammary carcinomas from animals pulse-labeled with BrdU were trisected. A portion was frozen and the remaining two portions were fixed in 10% neutral buffered formalin or methacarn for 24 hr, processed, embedded in paraffin, and sections stained for incorporated BrdU using a peroxidase immunohistochemical staining technique. Antigen retrieval techniques also were applied to formalin-fixed sections. Fixation in methacarn gave the highest labeling index (16.4%), which was comparable to that observed in unfixed frozen sections (17.5%). Formalin fixation alone dramatically suppressed the labeling index (0.3%), which was only partially recovered using various antigen retrieval techniques (2.1-8.1%). Methacarn fixation is recommended for prospective studies in which BrdU detection is planned because of the quantitative recovery of epitope and the simplicity of the approach.

scholarly journals Detection of Integrins in Formalin-fixed, Paraffin-embedded Tissues

1997 ◽  
Vol 45 (5) ◽  
pp. 737-741 ◽  
Author(s):  
Helen Liapis ◽  
Karen Hutton
Keyword(s):  
Tissue Expression ◽  
Enzyme Digestion ◽  
Optimal Conditions ◽  
Frozen Sections ◽  
Transmembrane Receptors ◽  
Archival Material ◽  
Formalin Fixed Paraffin ◽  
Formalin Fixed Paraffin Embedded ◽  
Formalin Fixed

Integrins are heterodimeric transmembrane receptors, which are expressed in many cells. In vitro experiments have demonstrated that integrins may be important in tumor progression and organ development. The functions of integrins were previously studied in cell cultures and their tissue expression was detected by immunofluorescence or immunoperoxidase in frozen sections. The purpose of this study was to determine the optimal conditions for detection of integrins in formalin-fixed, paraffin-embedded tissues. We utilized microwave heating and enzyme digestion in routinely processed, surgically removed tissues. Our results demonstrate that integrins can be reliably detected in archival material. This approach will facilitate further investigation of the role played by integrins in human malignancies and in developmental processes.

scholarly journals Antibodies validated for routinely processed tissues stain frozen sections unpredictably

BioTechniques ◽  
2021 ◽  
Vol 70 (3) ◽  
pp. 137-148
Author(s):  
Maddalena M Bolognesi ◽  
Francesco Mascadri ◽  
Laura Furia ◽  
Mario Faretta ◽  
Francesca M Bosisio ◽  
...  
Keyword(s):  
Frozen Sections ◽  
Formalin Fixed Paraffin ◽  
Nonspecific Staining ◽  
Formalin Fixed Paraffin Embedded ◽  
Antibody Validation ◽  
Formalin Fixed

Background: Antibody validation for tissue staining is required for reproducibility; criteria to ensure validity have been published recently. The majority of these recommendations imply the use of routinely processed (formalin-fixed, paraffin-embedded) tissue. Materials & methods: We applied to lightly fixed frozen sections a panel of 126 antibodies validated for formalin-fixed, paraffin-embedded tissue with extended criteria. Results: Less than 30% of the antibodies performed as expected with all fixations. 35% preferred one fixation over another, 13% gave nonspecific staining and 23% did not stain at all. Conclusion: Individual antibody variability of the paratope’s fitness for the fixed antigen may be the cause. Revalidation of established antibody panels is required when they are applied to sections whose fixation and processing are different from the tissue where they were initially validated.

scholarly journals THE VITAMIN A CONTENT AND RETINOL ESTERIFYING ACTIVITY OF A KUPFFER CELL FRACTION OF RAT LIVER

1972 ◽  
Vol 20 (10) ◽  
pp. 811-816 ◽  
Author(s):  
SAMUEL H. HORI ◽  
TAKAKO KITAMURA
Keyword(s):  
Vitamin A ◽  
Rat Liver ◽  
Kupffer Cell ◽  
Kupffer Cells ◽  
Qualitative Data ◽  
Fluorescence Microscope ◽  
Cell Fraction ◽  
Frozen Sections ◽  
Main Site ◽  
Formalin Fixed

Kupffer cells were isolated from normal and A-hypervitaminotic rats, and their vitamin A content and retinol esterifying activity were assayed in order to examine whether the Kupffer cells actually participate in the storage of vitamin A. The intrahepatic distribution of vitamin A was also studied using frozen sections of fresh and formalin-fixed livers by means of the fluorescence microscope. Both quantitative and qualitative data indicate that Kupffer cells are not the main site of vitamin A storage. The fluorescence of vitamin A and its reactivity to SbCl3 decreased drastically after formalin treatment.

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