Cloning of Tetrahymena cells in a chemically defined medium is possible in the presence of surfactants or at reduced temperature.

When Tetrahymena cells are exposed to physical or chemical stress they may die. The effect of a given stress depends on the culture medium, the temperature, and the manipulation of the cells. Cells in broth-medium or buffer solution are more resistant than cells in chemically defined medium (CDM).

Effects of Pluronic F-68 on Tetrahymena cells: protection against chemical and physical stress and prolongation of survival under toxic conditions.

The effects of the non-ionic surfactant Pluronic F-68 (0.01% w/v) on Tetrahymena cells have been studied. A marked protection against chemical and physical stress was observed.

Interface-mediated death of unconditioned Tetrahymena cells: effect of the medium composition.

We have previously shown that the cell death of Tetrahymena thermophila in low inocula cultures in a chemically-defined medium is not apoptotic. The death is caused by a cell lysis occurring at the medium-air interface and can be prevented by the addition of insulin or Pluronic F-68. Here, we report that cell death can also be caused by the medium.

Expression of Tetrahymena snRNA gene variants including a U1 gene with mutations in the 5' splice site recognition sequence.

The expression of U1, U2 and U5 snRNA gene variants has been studied under different physiological states of Tetrahymena. Variants of all three snRNA genes are expressed. Among the snRNAs detected is U1-3, a variant with 66 mutations compared to the normal U1 snRNA.

Surface mediated death of unconditioned Tetrahymena cells: effect of physical parameters, growth factors, hormones, and surfactants.

A new form of cell death has been observed. The death occurs at liquid-air interfaces when Tetrahymena cells are grown in a chemically defined medium (CDM) at low inocula. The cells die by lysis at the liquid-air interface (medium surface), which they reach due to negative gravitaxis as well as positive aerotaxis.

An improved quantitative assay for chemokinesis in Tetrahymena.

This paper presents a quantitative and sensitive assay for the measurement of chemosensory behavior in Tetrahymena. The two-phase assay is easy to perform in large quantities, so a variety of compounds can be screened under comparable conditions. A suspension of 2 x 10(5) cells ml-1 (the upper phase) is starved for 20-40 h and then gently placed on top of a 5% solution of Metrizamide (the lower phase) in a disposable microcuvette.

Physiological parameters affecting the chemosensory response of Tetrahymena.

We have investigated the significance of a number of physiological parameters in the preparation of cells for experiments on chemokinesis in Tetrahymena. The study comprises (1) growth state of the cells, (2) composition of the starvation medium, (3) concentration of cells during starvation, (4) oxygen saturation of the starvation medium, (5) temperature during starvation, and (6) starvation period. By controlling the physiological state of the cells, we significantly improved the reproducibility of the results obtained in assays for chemokinesis in Tetrahymena.

Characteristics of dividing and non-dividing Tetrahymena cells at different physiological states.

Eight defined physiological states of Tetrahymena pyriformis are described. For dividing cells the states comprise: 1. Exponentially growing cells, 2.

Certain fatty acids and steroids protect Tetrahymena from cell division stress caused by shaking.

Cultures of Tetrahymena are routinely shaken to ensure proper access to oxygen. Recent work showed that growth of dilute cultures (inocula < 10(4) cells ml-1) of T. pyriformis was sensitive to shaking.

Chemosensory behaviour of Tetrahymena.

Free swimming cells of the ciliated protozoan Tetrahymena are attracted to certain chemicals by chemokinesis. However, a special type of chemotaxis in response to a chemical gradient is found in cells gliding very slowly in semisolid media. In contrast to classical chemotaxis by leukocytes, which is solely positive towards chemo-attractants, the oriented chemokinesis by gliding Tetrahymena involves both positive and negative elements.

Chemotaxis in tetrahymena.

Chemotaxis - that is oriented locomotion of single cells - was shown by motion analysis of Tetrahymena thermophila. An electronic registration of swimming tracks was carried out in gradients of chemoattractant established in a modified Zigmond chamber. The attractants used were proteose peptone, platelet extract and fibroblast growth factor.

A paper membrane filter assay for ciliate chemoattraction.

A quantitative bioassay for ciliate chemoattraction based on the Boyden assay is described with the ciliates Tetrahymena thermophila and Tetrahymena pyriformis as test organisms. A chamber is separated into two compartments by a Whatman 3MM filter, and a suspension of starved cells (approximately 10(5) cells/ml) is placed in one compartment and a solution containing attractant in the other. The gradient of chemoattractant across the filter causes the cells to swim through the filter into the attractant-containing compartment where their appearance is determined by electronic cell counting.

Division competence in Tetrahymena: determination of minimum cell volume and rate of nutrient uptake.

Cell volume and doubling time have been determined for exponentially growing Tetrahymena pyriformis cells in broth medium with and without glucose and in media made from these media by dilution with water. The cells tolerate media with dry weights from 105 down to 0.06 g/L.

Cell volume and dry weight of cultured Tetrahymena.

Cultures of Tetrahymena pyriformis, T. hegewishi and T. malaccensis have been studied with regard to control of cell volume and cellular dry weight.

Immune hemolytic assay for identification of human anti-dsDNA antibodies with DNA-coated red blood cells as target cells.

A hemolytic assay was developed with the primary aim of being able to identify human lymphocytes producing anti-dsDNA antibodies found in patients with systemic lupus erythematosus (SLE). The coating of sheep red-blood cells with DNA was performed after precoating the cells with poly-L-lysine. The DNA-SRBC were lysed by anti-DNA antibodies from SLE sera, and the percent hemolysis was found to correlate with the anti-DNA activity demonstrated by the Farr assay (r = 0.

Parameters affecting the maximum cell concentration of Tetrahymena.

In cultures with efficient aeration a maximum cell concentration (MCC) of 6 X 10(5) cells/ml (defined medium) and 5.5 X 10(6) cells/ml (broth) can be reached. By culturing within Millicells with excess supply of medium and efficient removal of waste products a physical limit for MCC of about 13 X 10(6) cells/ml is reached.

Conjugation of DNA to erythrocytes.

DNA was conjugated to sheep red blood cells (SRBC) by chemical methods using CrCl3, poly-L-lysine or methylated bovine serum albumin as conjugation agents and by a physical method where conjugation was accomplished by incubation at 45 degrees C. The degree of conjugation was estimated using 32P-DNA (mean size 1 kbase pairs). Employing the CrCl3 method 5.

Small nuclear RNAs in the ciliate Tetrahymena.

We have isolated and partially characterized a family of small nuclear RNAs (snRNAs) from three different species of the protozoan Tetrahymena. We find six distinct snRNAs ranging in size from 100 to 250 nucleotides. The two largest snRNAs, as well as an abundant, heterogenous group of smaller snRNAs are found in the nucleolar RNA fraction.

Dimers of 5 S RNA and 5.8 S RNA in low molecular weight RNA preparations.

When purified 5 S RNA is exposed to concentrated solutions of sodium chloride or sodium phosphate, it is partly converted to the 5' S conformer but also partly to dimers and oligomers of 5 S RNA which are stable during gel filtration and electrophoresis. Presence of 5 S dimers can not be demonstrated in cellular RNA preparations extracted with phenol. 5.

Subcellular localization of low-molecular-weight RNA components in rat liver.

The subcellular localization of the four major low-molecular-weight RNA components, D, C, A and L, was studied in rat liver cells. The cells were fractionated by a non-aqueous technique into a nuclear and a cytoplasmic fraction. The cytoplasm contained 43% of component D, 57% of component C and more than 80% of component L.

Synthesis of low molecular weight RNA components in cells with a temperature-sensitive polymerase II.

AF 8 cells are a mutant cell line of baby hamster kidney cells with a temperature-sensitive polymerase II activity. When these cells grow at the non-permissive temperature (40 degrees C) the syntheseis of low molecular weight RNA components D, C and A is preferentially inhibited, whereas the synthesis of rRNA, tRNA, 5 S RNA and component L is affected only a little or not at all. These results indicate that polymerase II catalyzes the synthesis of components D, C and A.

Synthesis of low molecular weight RNA components A, C and D by polymerase II in alpha-amanitin-resistant hamster cells.

In an attempt to establish which RNA polymerase catalyzes the synthesis of the low molecular weight RNA components A, C and D, Ama 1 cells (mutant Chinese hamster cells) were used in experiments with addition of alpha-amanitin. Ama 1 cells contain an altered RNA polymerase II which is 800 times more resistant towards inhibition by alpha-amanitin than the wild type enzyme. Alpha-amanitin (up to 200 microgram/ml) added to these cells does not affect the synthesis of the low molecular weight RNAs A, C and D.