Doris Dahl

Localization of the glial fibrillary acidic protein in astrocytes by immunofluorescence

Abstract The glial fibrillary acidic (GFA) protein, a brain specific protein extracted from severely gliosed human tissue, is not species specific; cross-reaction occurs between anti-human GFA protein antibodies and brain extracts of rabbit, guinea pig, rat and dog. Using anti-GFA protein antiserum, astrocytes are selectively stained with the indirect immunofluorescence technique in both normal and pathological (gliosed) brain tissue.

THE ASTROGLIAL RESPONSE TO STABBING. IMMUNOFLUORESCENCE STUDIES WITH ANTIBODIES TO ASTROCYTE-SPECIFIC PROTEIN (GFA) IN MAMMALIAN AND SUBMAMMALIAN VERTEBRATES

The astroglial response t o stabbing. Immunofluorescence studies with antibodies to astrocyte‐specific protein (GFA) in mammalian and sub‐mammalian vertebrates

Localization of vimentin, the nonspecific intermediate filament protein, in embryonal glia and in early differentiating neurons: In vivo and in vitro immunofluorescence study of the rat embryo with vimentin and neurofilament antisera

Abstract Antisera raised against vimentin, the protein subunit of nonspecific intermediate-sized filaments (IFs), were used in conjunction with neurofilament (NF) antisera to study the early development of neurons and glia in the rat embryo. Vimentin-positive fibers spanning the entire thickness of the neural tube including the cerebral vesicles were first observed on Day 12, concomitant with the appearance of NF protein in more confined areas (anterolateral regions of spinal cord and brain stem; motor roots emerging from the NF-positive areas). From Day 15 onwards vimentin and NF antisera selectively decorated glia and neurons, respectively, both in vivo and in vitro . Before Day 15 it appeared that NF-positive structures also stained with antivimentin in cryostat sections. In vitro experiments confirmed the presence of vimentin in early differentiating neurons. NF-positive cells were observed which also reacted with antivimentin in cultures obtained from 13- and 14-day embryos, but not later in development. Most neurons in these cultures became vimentin negative after 2–3 days in vitro .

The radial glia of Müller in the rat retina and their response to injury. An immunofluorescence study with antibodies to the glial fibrillary acidic (GFA) protein

Abstract The glial response to injury was studied in the rat retina by immunofluorescence with GFA antisera. In the normal rat only the glia limitans on the inner surface of the retina was positive by immunofluorescence. Following interruption of the optic nerve or multiple penetrating wounds of the eye the radial fibres of Muller became intensely stained. It is concluded that Muller glia are similar to brain astrocytes with respect to their reaction to injury, that is they accumulate GFA protein.

Laminin is induced in astrocytes of adult brain by injury.

Laminin is a high mol. wt. non‐collagenous matrix glycoprotein, confined in adult tissues to basement membranes. In normal rat brain we found laminin mainly in vessel walls but, after injury, induced by stereotaxic injection of a neurotoxin, laminin immunoreactivity appeared also in reactive astrocytes, which are characteristically positive for the glial fibrillary acidic protein (GFAP). Laminin was first detected in GFAP‐immunoreactive glial cells 24 h after injury. Four days later the majority of reactive astrocytes in the gray matter were positive for laminin and the laminin immunoreactivity, but not that of GFAP, gradually subsided within a month. Fibronectin, the other major matrix glycoprotein, was found only in capillary structures both in normal and lesioned brain tissue. The results indicate that mature astrocytes have the potential to produce laminin and suggest a role for this glycoprotein in brain regeneration.

Differentiation of astrocytes in the cerebellar cortex and the pyramidal tracts of the newborn rat. An immunofluorescence study with antibodies to a protein specific to astrocytes

An automatic machine for assembling and banding together the components making up an expansion shell so that the assembly can be conveniently packaged and shipped. In one embodiment, the machine contains a series of work stations arranged to assemble the bolt components within a die. The expansion shell halves are staked to a common strap and the components then passed through a combination forming gage and banding mechanism which aligns the parts in assembly and bands them together using a plastic sleeve. In a second embodiment of the invention, the expansion shells are staked to the strap prior to being loaded into the die.

Immunohistochemical localization of glial fibrillary acidic protein in human glial neoplasms

The presence and distribution of glial fibrillary acidic protein in fixed, paraffin embedded tissue were studied in 85 human intracranial neoplasms, using the peroxidase‐anti‐peroxidase method. In some cases, indirect immunofluorescence of frozen sections was used as well. In normal tissue, only the cell processes and perikarya of fibrous astrocytes were stained. Immunostaining was also observed in the following glial neoplasms: astrocytomas (all varieties), astroblastoma, subependymal giant cell astrocytoma, subependymoma, glioblastoma multiforme and ependymoma. The astrocytic elements of mixed gliomas and of medulloblastomas undergoing glial differentiation were likewise strongly stained. In contrast, oligodendrogliomas, meningiomas, pituitary adenomas, sarcomas, lymphomas and metastatic carcinomas were negative. Either a perikaryal or a diffuse fibrillary staining pattern was observed. Combination of the two patterns occasionally occurred. The perikaryal staining was prominent in gemistocytic astrocytomas and in astroblastomas. A distinct negative correlation existed between the degree of anaplasia and the intensity of immunostaining.

Specificity of the glial fibrillary acidic protein for astroglia.

Glial fibrillary acidic protein (GFA) is the main constituent of glial filaments and the close similarity of GFA and neurofilament protein has been recently reported. However, the immunofluorescence staining of peripheral nerve which may be observed with GFA antisera is not due to cross-reaction between GFA and neurofilament protein. Staining of peripheral axons was also observed with control sera obtained by injecting the rabbits with nonimmunogenic GFA preparations isolated with the same procedure. Immune GFA antisera and control sera reacted with sodium dodecyl sulfate extracts of sciatic nerve. However, the precipitin line formed with peripheral nerve crossed the line against GFA protein, thus indicating nonidentity between the two antigens. Buffer extract of sciatic nerves that had been incubated with spinal cord reacted by immunodiffusion with GFA antisera, thus indicating that redistribution of GFA occurred under these conditions.

Glial Fibrillary Acidic Protein (GFA) in Normal Neural Cells and in Pathological Conditions

Publisher Summary This chapter discusses glial fibrillary acidic (GFA) protein in normal neural cells and in pathological conditions. A review of the literature on GFA protein indicates that it has found its major application as an immunohistochemical marker for astroglia, especially under conditions where identification of the cell type is difficult, that is, during development, in culture, in cellular fractions and, more recently, in human brain tumors. Laboratory procedures have been developed to facilitate production of the antisera. The solubility of GFA protein in aqueous solutions and the in vitro formation of filaments from purified GFA protein preparations suggest that glial filaments, like other constituents of the cytoskeleton in nonmuscle cells, namely, microtubules and actin microfilaments, may assemble and disassemble in the cytoplasm. The extensive remodeling of the glial framework during brain development and the rapid change in cell shape occurring in cultured astrocytes following serum withdrawal and the administration of cyclic AMP make this hypothesis particularly attractive. The original isolation of GFA protein from gliosed human brain required relatively simple procedures such as buffer extraction and ammonium sulfate precipitation to obtain purified preparations. The immunogenicity of GFA protein varies considerably depending on the degree of degradation. Nondegraded or partially degraded preparations are nonimmunogenic or weakly immunogenic whereas strong antisera are obtained with degraded antigen. Immunohistological studies with GFA protein antisera provide new data in three critical areas of research on the development of neuroglia: (1) myelination gliosis, (2) time of appearance of Bergmann fibers in the cerebellum, and (3) the nature of radial glia.

Immunochemical and immunofluorescence studies of the glial fibrillary acidic protein in vertebrates

Abstract Phosphate buffer extracts (pH 8.0) of the brain and the spinal cord of various vertebrates, mammalian and submammalian, cross-reacted with antihuman glial fibrillary acidic (GFA) protein serum by Ouchterlony double diffusion. Mammalian brain extracts, as previously reported, gave an immunodiffusion pattern of complete identity with human GFA protein. In submammalian vertebrates (shark, goldfish, turtle and chicken) strong spurs were observed at the junction with human GFA protein, suggesting incomplete identity. GFA protein from mammalian and submammalian vertebrates had the same extraction properties, immunoelectrophoretic mobility and molecular weight as human GFA protein. By immunofluorescence with antihuman GFA protein, serum astrocytes were selectively stained in cryostat sections of the brain and the spinal cord in all vertebrates studied, mammalian and submammalian, with the exception of the lamprey. No immunofluorescence was observed in invertebrate ganglia. Some antomical observations on the astrocytic framework of various vertebrates, as observed with immunofluorescence, are reported.