Formalin inactivation of the lactate dehydrogenase-elevating virus reveals a major neutralizing epitope not recognized during natural infection.

Five hybridomas that secrete monoclonal antibodies which neutralize the infectivity of lactate dehydrogenase-elevating virus (LDV) were isolated from BALB/c mice primed with Formalin-inactivated LDV. Competition analyses indicated that all five neutralizing monoclonal antibodies recognize contiguous, if not identical, epitopes on the envelope glycoprotein of LDV (VP-3) which are not recognized by nonneutralizing VP-3-specific monoclonal antibodies isolated from the same fusion. Despite the presence of neutralizing activity, polyclonal anti-LDV antibodies obtained from persistently infected mice did not compete for binding to LDV with four of the five neutralizing monoclonal antibodies tested. The results indicate that the envelope glycoprotein of LDV possesses a major neutralizing epitope which is poorly recognized, if at all, by mice during a natural infection but is rendered immunogenic by Formalin inactivation of the virus. The epitope was also not immunogenic in a rabbit, since its polyclonal LDV-neutralizing antibodies did not inhibit binding of the mouse monoclonal antibodies to LDV. Passive immunization with the neutralizing monoclonal antibodies did not protect mice from LDV infection and did not alter the course of infection. Neutralizing monoclonal antibodies have been used to select a neutralization escape variant by a novel combination of in vitro and in vivo isolation.     

Monoclonal antibody protection from age-dependent poliomyelitis: implications regarding the pathogenesis of lactate dehydrogenase-elevating virus.

Over 90% of cyclophosphamide-treated, 6- to 7-month-old C58/M mice developed fatal paralytic disease after infection with a virulent strain of lactate dehydrogenase-elevating virus (LDV), with a mean onset of paralysis of about 16 days. Passive immunization with polyclonal antibodies or with a group of anti-LDV monoclonal antibodies (MAbs) with single-epitope specificity 1 day before or at the time of LDV infection prevented the development of paralytic disease without interfering with the replication of LDV in permissive macrophages, the primary host cells of LDV. In situ hybridization of spinal cord sections with an LDV-specific cDNA probe indicated that the MAb specifically prevented the cytocidal infection of motor neurons by LDV without blocking the infection of smaller nonneuronal cells in the spinal cord. The protective antibodies recognize at least two different epitopes on the glycoprotein of LDV, VP-3. Passive immunizations with other anti-LDV MAbs, which recognize at least three other epitopes on VP-3 of LDV, afforded no protection. In contrast to the protective effect of anti-LDV MAb injection before or at the time of LDV infection, their administration postinfection exerted relatively little protection, though it delayed the appearance of paralytic symptoms. However, repeated injections of MAbs until at least 7 days postinfection also afforded a high degree of protection. The results indicate that protective MAbs may interfere with two stages in the development of LDV-induced paralytic disease. When administered at the time of LDV infection, they prevent the initial infection of spinal cord motor neurons. After this initial event, repeated injections of MAb are required to inhibit the spread of LDV between neurons until the endogenous production of protective anti-LDV antibodies in these mice.     

Cytohistologic correlation of urothelial lesions secondary to photodynamic therapy

Qualitative cytologic evaluations of urinary bladder washings were performed on a selected population following photodynamic therapy for recurrent transitional cell carcinoma of the bladder. The seven patients were monitored trimonthly by cystoscopy, multiple biopsies and cytopreparations. Cancers reappeared in two of the five patients who initially responded to therapy. In the remaining two patients, the recurrent neoplasms were therapeutically refractory. Cytology detected recurrent cancer prior to biopsy confirmation and/or cytoscopic identification. Exfoliative cytology was correlated with the histopathology of the concurrent biopsies; a possible source for a false-positive cytodiagnosis was the cellular atypia of reepithelialized bladder mucosa. Dysplasia was not identified cytologically or histologically.     

Phylogenetic diversification of immunoglobulin genes and the antibody repertoire

Immunoglobulins are encoded by a large multigene system that undergoes somatic rearrangement and additional genetic change during the development of immunoglobulin-producing cells. Inducible antibody and antibody-like responses are found in all vertebrates. However, immunoglobulin possessing disulfide-bonded heavy and light chains and domain-type organization has been described only in representatives of the jawed vertebrates. High degrees of nucleotide and predicted amino acid sequence identity are evident when the segmental elements that constitute the immunoglobulin gene loci in phylogenetically divergent vertebrates are compared. However, the organization of gene loci and the manner in which the independent elements recombine (and diversify) vary markedly among different taxa. One striking pattern of gene organization is the "cluster type" that appears to be restricted to the chondrichthyes (cartilaginous fishes) and limits segmental rearrangement to closely linked elements. This type of gene organization is associated with both heavy- and light-chain gene loci. In some cases, the clusters are "joined" or "partially joined" in the germ line, in effect predetermining or partially predetermining, respectively, the encoded specificities (the assumption being that these are expressed) of the individual loci. By relating the sequences of transcribed gene products to their respective germ-line genes, it is evident that, in some cases, joined-type genes are expressed. This raises a question about the existence and/or nature of allelic exclusion in these species. The extensive variation in gene organization found throughout the vertebrate species may relate directly to the role of intersegmental (V<==>D<==>J) distances in the commitment of the individual antibody-producing cell to a particular genetic specificity. Thus, the evolution of this locus, perhaps more so than that of others, may reflect the interrelationships between genetic organization and function.      

Action Spectra for Conversions of Phycochrome c from Nostoc muscorum

The reversibly photochromic pigment, phycochrome c, was extracted from the blue-green alga Nostoc muscorum strain A. Action spectra were determined for in vitro conversions of the pigment from the short wavelength to the long wavelength form and vice versa. The action peak for the absorbance decrease at 650 nm is at 630 nm. During this decrease there is only a slight increase of the absorbance in the green region. Green and yellow light (maximum efficiency at 580 nm) completely restores absorbance at 650 nm. The observations are explained by the existence of three spectrally different forms of phycochrome c: P^c₆₃₀ and P^c₆₅₀ which equilibrate in darkness and P^c₅₈₀ which is reversibly photoconvertible to P^c₆₃₀. We have also measured the absorbance changes brought about by saturating irradiations with light of various wavelengths (“photostationary state spectrum”). Extreme photostationary states were obtained with about 650 nm and 500 nm light.     

Effects of Frost Hardening and Dehardening on Photosynthetic Electron Transport and Fluorescence Properties in Isolated Chloroplasts of Pinus silvestris

Photosynthetic electron transport and low-temperature fluorescence emission properties have been analyzed in isolated chloroplasts during the course of frost hardening and dehardening of Pinus silvestris L. Both the partial electron-transport reactions (H₂O DPIP and Asc./DPIP NADP) and the overall electron transport (H₂O — NAPD) showed decreasing capacities during the course of hardening. Upon exposing the plants to ?5°C and high irradiance a block in the electron-transport chain between the two photosystems developed, whereas the partial reactions still showed activities. The decrease in activity of PSl was accompanied by a decrease in P700 content, as determined by light oxidation of P700, which indicates a correlation between the two changes. Hardening also induced changes in the in vivo chlorophyll organization. During the course of hardening the fluorescence emission bands F692 and F726 decreased relative to F680. These changes were more pronounced if the plants were treated in high than in low irradiance. This suggests a greater destruction of the chlorophyll antennae in close association with the two photoreactions than in the so-called light-harvesting chlorophyll a/b antenna. During dehardening basically the reverse of the changes observed during hardening occurred. The recovery of secondary needles was complete, whereas primary needles only partly recovered.