scholarly journals In vitro assay for erythropoietin: erythroid colony formation in methyl cellulose used for the measurement of erythropoietin in plasma

Blood ◽  
1979 ◽  
Vol 53 (6) ◽  
pp. 1172-1181 ◽  
Author(s):  
P Haga ◽  
B Falkanger
Keyword(s):  
Colony Formation ◽  
Methyl Cellulose ◽  
High Sensitivity ◽  
In Vitro Assay ◽  
Target Cells ◽  
Newborn Mouse ◽  
Newborn Mice ◽  
Vitro Assay ◽  
Good Agreement

Abstract Erythroid colony formation in methyl cellulose has been used for the measurement of erythropoietin in plasma. Livers from newborn mice less than 24 hr old were found to provide convenient target cells. Newborn mouse liver contains a substantial number of erythroid colony-forming cells (CFU-e) that have a high sensitivity to erythropoietin, the dose-- response curve for erythropoietin reaching a plateau at 50 mU/ml. As little as 0.5 m/ml of the hormone is detectable. Removal of cells that adhered to glass prior to culturing doubled the number of colonies formed in the presence of erythropoietin. Addition of untreated plasmas that showed high erythropoietin titers in the exhypoxic polycythemic mice assay gave variable results. Some of the plasmas stimulated colony formation actively and in a linear fashion. However, the majority of the plasmas were toxic to the cultures. Dialyzing the plasmas for 3 days against distilled water effectively removed the toxicity. Results obtained with the method are in good agreement with the values found using the exhypoxic polycythemic mice assay.

scholarly journals In vitro assay for erythropoietin: erythroid colony formation in methyl cellulose used for the measurement of erythropoietin in plasma

Blood ◽  
1979 ◽  
Vol 53 (6) ◽  
pp. 1172-1181 ◽  
Author(s):  
P Haga ◽  
B Falkanger
Keyword(s):  
Colony Formation ◽  
Methyl Cellulose ◽  
High Sensitivity ◽  
In Vitro Assay ◽  
Target Cells ◽  
Newborn Mouse ◽  
Newborn Mice ◽  
Vitro Assay ◽  
Good Agreement

Erythroid colony formation in methyl cellulose has been used for the measurement of erythropoietin in plasma. Livers from newborn mice less than 24 hr old were found to provide convenient target cells. Newborn mouse liver contains a substantial number of erythroid colony-forming cells (CFU-e) that have a high sensitivity to erythropoietin, the dose-- response curve for erythropoietin reaching a plateau at 50 mU/ml. As little as 0.5 m/ml of the hormone is detectable. Removal of cells that adhered to glass prior to culturing doubled the number of colonies formed in the presence of erythropoietin. Addition of untreated plasmas that showed high erythropoietin titers in the exhypoxic polycythemic mice assay gave variable results. Some of the plasmas stimulated colony formation actively and in a linear fashion. However, the majority of the plasmas were toxic to the cultures. Dialyzing the plasmas for 3 days against distilled water effectively removed the toxicity. Results obtained with the method are in good agreement with the values found using the exhypoxic polycythemic mice assay.

scholarly journals Demonstration of burst-promoting activity of recombinant human GM-CSF on circulating erythroid progenitors using an assay involving the delayed addition of erythropoietin

Blood ◽  
1985 ◽  
Vol 66 (6) ◽  
pp. 1479-1481 ◽  
Author(s):  
RE Donahue ◽  
SG Emerson ◽  
EA Wang ◽  
GG Wong ◽  
SC Clark ◽  
...  
Keyword(s):  
T Cell ◽  
Cell Line ◽  
Peripheral Blood ◽  
Colony Formation ◽  
In Vitro Assay ◽  
Erythroid Progenitors ◽  
Vitro Assay ◽  
Gm Csf ◽  
Semi Solid

Abstract We demonstrate through the use of an in vitro assay involving the delayed addition of erythropoietin that human recombinant GM-CSF, cloned from a mature T cell line, Mo, clearly has burst-promoting activity (BPA) on peripheral blood erythroid progenitors at picomolar concentrations. Delay for up to 72 hours of the addition of erythropoietin to semi-solid methylcellulose cultures of concentrated peripheral blood progenitors minimizes or eliminates BPA-independent erythroid colony formation with little loss of BPA-dependent erythroid colony formation. This assay will prove useful in accurately detecting sources of BPA.

COLONY FORMATION IN AGAR: IN VITRO ASSAY FOR HAEMOPOIETIC STEM CELLS

1971 ◽  
Vol 4 (5) ◽  
pp. 463-477 ◽  
Author(s):  
K. A. Dicke ◽  
M. G. C. Platenburg ◽  
D. W. Bekkum
Keyword(s):  
Stem Cells ◽  
Colony Formation ◽  
In Vitro Assay ◽  
Vitro Assay ◽  
Haemopoietic Stem Cells

scholarly journals Demonstration of burst-promoting activity of recombinant human GM-CSF on circulating erythroid progenitors using an assay involving the delayed addition of erythropoietin

Blood ◽  
1985 ◽  
Vol 66 (6) ◽  
pp. 1479-1481 ◽  
Author(s):  
RE Donahue ◽  
SG Emerson ◽  
EA Wang ◽  
GG Wong ◽  
SC Clark ◽  
...  
Keyword(s):  
T Cell ◽  
Cell Line ◽  
Peripheral Blood ◽  
Colony Formation ◽  
In Vitro Assay ◽  
Erythroid Progenitors ◽  
Vitro Assay ◽  
Gm Csf ◽  
Semi Solid

We demonstrate through the use of an in vitro assay involving the delayed addition of erythropoietin that human recombinant GM-CSF, cloned from a mature T cell line, Mo, clearly has burst-promoting activity (BPA) on peripheral blood erythroid progenitors at picomolar concentrations. Delay for up to 72 hours of the addition of erythropoietin to semi-solid methylcellulose cultures of concentrated peripheral blood progenitors minimizes or eliminates BPA-independent erythroid colony formation with little loss of BPA-dependent erythroid colony formation. This assay will prove useful in accurately detecting sources of BPA.

In Vitro Assay of Platelet Adhesiveness with Washed and Tanned Human Red Cells

1968 ◽  
Vol 20 (03/04) ◽  
pp. 384-396 ◽  
Author(s):  
G Zbinden ◽  
S Tomlin
Keyword(s):  
Platelet Adhesion ◽  
Sodium Citrate ◽  
Blood Platelets ◽  
In Vitro Assay ◽  
Red Cells ◽  
Platelet Adhesiveness ◽  
Vitro Assay ◽  
In Vitro System ◽  
Human Red Cells

SummaryAn in vitro system is described in which adhesion of blood platelets to washed and tannic acid-treated red cells was assayed quantitatively by microscopic observation. ADP, epinephrine and TAME produced a reversible increase in platelet adhesiveness which was antagonized by AMP. With Evans blue, polyanetholsulfonate, phthalanilide NSC 38280, thrombin and heparin at concentrations above 1-4 u/ml the increase was irreversible. The ADP-induced increase in adhesiveness was inhibited by sodium citrate, EDTA, AMP, ATP and N-ethylmaleimide. EDTA, AMP and the SH-blocker N-ethylmaleimide also reduced spontaneous platelet adhesion to red cells. No significant effects were observed with adenosine, phenprocoumon, 5-HT, phthalanilide NSC 57155, various estrogens, progestogens and fatty acids, acetylsalicylic acid and similarly acting agents, hydroxylamine, glucose and KCN. The method may be useful for the screening of thrombogenic and antithrombotic properties of drugs.

scholarly journals Validation of the in vitro comet assay for DNA cross-links and altered bases detection

2021 ◽  
Author(s):  
Damián Muruzabal ◽  
Julen Sanz-Serrano ◽  
Sylvie Sauvaigo ◽  
Bertrand Treillard ◽  
Ann-Karin Olsen ◽  
...  
Keyword(s):  
Comet Assay ◽  
Human Health Risk ◽  
High Sensitivity ◽  
Dna Lesions ◽  
Dna Glycosylase ◽  
Vitro Assay ◽  
Cytotoxic Compound ◽  
Endonuclease Iii ◽  
Cross Links

AbstractMechanistic toxicology is gaining weight for human health risk assessment. Different mechanistic assays are available, such as the comet assay, which detects DNA damage at the level of individual cells. However, the conventional alkaline version only detects strand breaks and alkali-labile sites. We have validated two modifications of the in vitro assay to generate mechanistic information: (1) use of DNA-repair enzymes (i.e., formamidopyrimidine DNA glycosylase, endonuclease III, human 8-oxoguanine DNA glycosylase I and human alkyladenine DNA glycosylase) for detection of oxidized and alkylated bases as well as (2) a modification for detecting cross-links. Seven genotoxicants with different mechanisms of action (potassium bromate, methyl methanesulfonate, ethyl methanesulfonate, hydrogen peroxide, cisplatin, mitomycin C, and benzo[a]pyrene diol epoxide), as well as a non-genotoxic compound (dimethyl sulfoxide) and a cytotoxic compound (Triton X-100) were tested on TK-6 cells. We were able to detect with high sensitivity and clearly differentiate oxidizing, alkylating and cross-linking agents. These modifications of the comet assay significantly increase its sensitivity and its specificity towards DNA lesions, providing mechanistic information regarding the type of damage.

Angiogenic Effect of Pravastatin Alone and with Sera from Healthy and Complicated Pregnancies Studied by in vitro Vasculogenesis/Angiogenesis Assay

2021 ◽  
pp. 1-9
Author(s):  
Anita Virtanen ◽  
Outi Huttala ◽  
Kati Tihtonen ◽  
Tarja Toimela ◽  
Tuula Heinonen ◽  
...  
Keyword(s):  
Direct Effect ◽  
Late Onset ◽  
Maternal Serum ◽  
In Vitro Assay ◽  
Serum Samples ◽  
Therapeutic Concentration ◽  
Tubule Formation ◽  
Vitro Assay ◽  
Angiogenesis Assay

<b><i>Objective:</i></b> To determine the direct effect of pravastatin on angiogenesis and to study the interaction between pravastatin and maternal sera from women with early- or late-onset pre-eclampsia (PE), intrauterine growth restriction, or healthy pregnancy. <b><i>Methods:</i></b> We collected 5 maternal serum samples from each group. The effect of pravastatin on angiogenesis was assessed with and without maternal sera by quantifying tubule formation in a human-based in vitro assay. Pravastatin was added at 20, 1,000, and 8,000 ng/mL concentrations. Concentrations of angiogenic and inflammatory biomarkers in serum and in test medium after supplementation of serum alone and with pravastatin (1,000 ng/mL) were measured. <b><i>Results:</i></b> Therapeutic concentration of pravastatin (20 ng/mL) did not have significant direct effect on angiogenesis, but the highest concentrations inhibited angiogenesis. Pravastatin did not change the levels of biomarkers in the test media. There were no changes in angiogenesis when therapeutic dose of pravastatin was added with maternal sera, but there was a trend to wide individual variation towards enhanced angiogenesis, particularly in the early-onset PE group. <b><i>Conclusions:</i></b> At therapeutic concentration, pravastatin alone or with maternal sera has no significant effect on angiogenesis, but at high concentrations the effect seems to be anti-angiogenic estimated by in vitro assay.

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