In vitro and in vivo inhibition of monocyte phagocytic function by factor VIII concentrates: correlation with concentrate purity

SummaryIn response to reports of discrepant in vitro assays of high-purity concentrates, a double-blind crossover study of in vivo recovery and half-life of two brands of monoclonal-antibody-purified factor VIII concentrates (Monoclate and Hemofil-M) was performed in 23 patients with hemophilia A. In vivo recoveries were close to values predicted from the labelled unitage when plasma samples were assayed by a one-stage method. When a two-stage assay was used, lower recoveries were calculated and the recovery with Hemofil-M was slightly but significantly lower than that with Monoclate. The concentrates were re-assayed in vitro by the two-stage method. Monoclate (which is assayed by the manufacturer using a two-stage method) contained 97% of the labelled potency and Hemofil-M (which is assayed by the manufacturer using a one-stage method) contained 81% of the labelled potency. Differences in in vitro and in vivo assay methods contribute to disparities between expected and observed factor VIII recovery. Clearance of Hemofil-M was significantly faster than that of Monoclate, but volume of distribution at the steady state, mean residence time, and plasma half-disappearance times of the two concentrates were not significantly different.

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Human Factor VIII Concentrates in Hemophiliac Dogs

10.1055/s-0039-1682593 ◽

1977 ◽

Author(s):

Jessica H. Lewis ◽

Ute Hasiba ◽

Joel A. Spero

Keyword(s):

Factor Viii ◽

Human Factor ◽

Rapid Development ◽

The Third ◽

Human Factor Viii ◽

Post Infusion ◽

Factor Viii Concentrates ◽

In Vitro Effects

Human factor VIII corrects the clotting defect in dog hemophilic plasma in vitro. The present studies were undertaken to see if this happened in vivo and to look for and document the development of an inhibitor. Four hemophiliac dogs were infused with factor VIII concentrates, the first two on five occasions, the others three times. Factor VIII:C, VIIIR:Ag (defined with antibody to human VIII) and VIIIR:vW were followed at pre, 10 minutes, 2 and 24 hours post infusion. The pre-infusion VIII:C (assayed with human substrate) averaged 0.23 U/ml compared to 6.93 U/ml for normal dogs; VIIIR:Ag was absent in both. VIIIR:vW was low but variable. Following the first injection, all four dogs responded in VIII:C about as calculated. The amounts of VIIIR:Ag and vW were much greater than VIII:C in the concentrates and in the post-first infusion samples from the dogs. On subsequent infusions rises in VIIIR:Ag were not detected and increases in VIII:C and VIIIR:vW were minimal. Precipitating anti-human VIII was found on the third infusion and thereafter. After the first infusion reactions were marked. Vomiting and diarrhea occurred in all, and one dog died in anaphylactic shock about one hour after the third infusion. Lack of response in VIIIR:Ag occurred before anti-VIII could be demonstrated in vitro. This rapid development of an inhibitor suggests that prolonged cross-species VIII therapy will not be successful. The ability of the precipitating anti-VIII elicited in the dogs to destroy VIII:C, VIIIR:Ag and VIIIR:vW is analagous to the in vitro effects of heterologous anti-VIIIs (rabbit and goat).

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THE PRACTICAL VALUE OF THE IN VITRO FILTER BLEEDING TIME

10.1055/s-0038-1642847 ◽

1987 ◽

Author(s):

G P Salmon ◽

J R O’Brien

Keyword(s):

Factor Viii ◽

Bleeding Time ◽

Normal Range ◽

Normal Test ◽

Factor Viii Concentrates ◽

Von Willebrand ◽

Sample Coefficient Of Variation ◽

Test Treatment

The in vivo bleeding time has many disadvantages, some of which are obviated by an in vitro filter bleeding time. Normal native or heparinized blood is forced at 40mmHg through a filter made of glass microfibres with tortuous capillary sized pores which retain particles over lOμ. It is found that with high shear platelet aggregation occurs and when about 200,000 platelets (or 70/O of platelets are retained the filter becomes blocked. The test takes about two minutes to perform. Three routine measurements are easily made. (1) The % platelets retained between 10-20 secs; (2) the rate of blocking, e.g. drops 10-20/drops 0-5; (3) the blocking time. The within sample coefficient of variation (c of v) was 9% ± 4, n = 6. The c of v within stable patients studied over six months was 20% ± 11, n = 20. The correlation between platelet retention and the skin bleeding time is good overall when von Willebrand (VW) patients are included (r = ™0.73, n = 52) but absent when both tests are within the normal range. This test is sensitive to R:ag values from 0-160 (r = 0.62,n = 52) but poor within the VW and within normal groups. It is also abnormal in some patients with low platelet counts. Atherosclero-tics and diabetics have a normal test. Treatment of VW patients with DDAVP or with cryoprecipitate usually normalises this test. In one VW patient neither substance corrected this test and he bled. The test is thus useful in monitoring in vitro and in vivo the efficiency of various factor VIII concentrates to stop bleeding in VW disease. In vitro it is abnormal in the presence of many “membrane-active” drugs. Different drugs also have different effects on the retention of white cells; so it could be useful as a pharmacological screen. Thus it has uses as a quick and easy clinical screen for many forms of platelet-dependent haemostatic defects. It could also be used by the pharmaceutical industry and by producers of factor VIII concentrates.

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Issues with the Assay of Factor VIII Activity in Plasma and Factor VIII Concentrates

Thrombosis and Haemostasis ◽

10.1055/s-0037-1614153 ◽

2000 ◽

Vol 84 (12) ◽

pp. 942-948 ◽

Cited By ~ 28

Author(s):

Henry Kingdon ◽

Kenneth Mann ◽

Gilbert White ◽

Roger Lundblad

Keyword(s):

Factor Viii ◽

Hemophilia A ◽

Coagulation Factors ◽

Chromogenic Substrate ◽

In Vitro Measurements ◽

Factor Viii Concentrates ◽

The One

SummaryA review of the literature suggests that assays accurate for the determination of factor VIII in plasma samples may not necessarily retain this accuracy when used for the determination of factor VIII in high-purity factor VIII concentrates such as Hemofil ® M. Review of assay data suggests that it is imperative to obtain maximal activation of the factor VIII in the sample with thrombin when using an assay system of isolated coagulation factors such as the two-stage assay or the various chromogenic substrate assays. Based on a combination of ease and reproducibility of performance and correlation of in vivo and in vitro measurements, it is recommended that the one-stage activated partial thromboplastin time performed with plasma from an individual with severe hemophilia A be used for the measurement of factor VIII potency. Chromogenic substrate assays can be used if care is taken to assure optimal activation of factor VIII by thrombin in the assay and the presence of sufficient factor IXa, phospholipid and calcium ions to stabilize factor Villa during the assay process.

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Decreased Potency Of Factor VIII Concentrates

10.1055/s-0038-1652637 ◽

1981 ◽

Cited By ~ 3

Author(s):

C K Kasper

Keyword(s):

Factor Viii ◽

In Vitro Assays ◽

Assay Method ◽

Two Stage ◽

One Stage ◽

Plasma Factor ◽

Factor Viii Concentrates ◽

The One

Plasma factor VIII recoveries after infusions of factor VIII concentrates into patients with classic hemophilia have been measured in this laboratory for 14 years. Recently, we observed a decline in the in vivo recovery of factor VIII per factor VIII unit infused. In 1980, plasma factor VIII levels were measured by a one-stage APTT-based assay before and 10 min after 150 infusions of 46 lots of 3 brands of factor VIII concentrate produced in the U.S.A. Our pooled normal plasma reference was calibrated against WHO International Standard 2 and results expressed in International factor VIII units. Observed in vivo factor VIII recovery was compared to the value expected from calculations based on the unitage stated on the label. The ratio of observed/expected recovery averaged 56% per lot for brand A, 60% per lot for brand B, and 103% per lot for brand C. In vitro assays were performed on 22 lots on 36 occasions, and the ratio of observed/labelled units average 46% per lot for brand A, 53% for brand B and 75% for brand C. The two-stage factor VIII assay method of Pool and Robinson was also used to assay plasma samples from 18 infusions, and results averaged 135% of the one-stage values for infusions of brand A, 160% for brand B, and 109% for brand C. (Brand A is assayed by the manufacturer by a two-stage method, brands B and C by one-stage methods.)Decreased clinical efficacy was observed when postinfusion plasma factor VIII levels were lower than customary. The decline in potency of brands A and B has necessitated more frequent assay of patients and use of larger amounts of concentrate, with greatly-increased expense. Investigation of the effect of different assay methods and different factor VIII standards and references on the apparent factor VIII content of concentrates has begun.

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In Vitro and in Vivo Evaluation of Two Factor VIII Concentrates Virally Inactivated by Solvent-Detergent or by Pasteurization

Acta Clinica Belgica ◽

10.1080/17843286.1991.11718180 ◽

1991 ◽

Vol 46 (5) ◽

pp. 298-304 ◽

Cited By ~ 2

Author(s):

K. Peerlinck ◽

J. Amout ◽

M. Tamise ◽

A. Vanherle ◽

P. Fondu ◽

...

Keyword(s):

Factor Viii ◽

In Vivo Evaluation ◽

Factor Viii Concentrates

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Specific factor VIII-related antigen fragmentation: an in vivo and in vitro phenomenon

Blood ◽

10.1182/blood.v60.4.930.930 ◽

1982 ◽

Vol 60 (4) ◽

pp. 930-939 ◽

Cited By ~ 14

Author(s):

RR Montgomery ◽

J Johnson

Keyword(s):

Factor Viii ◽

Bleeding Time ◽

Antigenic Determinants ◽

Specific Factor ◽

Crossed Immunoelectrophoresis ◽

Agarose Electrophoresis ◽

Factor Viii Concentrates

Abstract Factor VIII concentrates have been shown to have reduced ability to correct the bleeding time defect in von Willebrand's disease and to have abnormal mobility of their VIIIR:Ag on crossed immunoelectrophoresis. This report concerns the partial characterization of a fragment of VIIIR:Ag that lacks some of the normal antigenic determinants present on VIIIR:Ag. It is present in commercial factor VIII concentrates, but not in cryoprecipitate, and is recovered from the plasma of hemophilic patients who have been infused with these concentrates. The fragment is also produced during disseminated intravascular coagulation. Although it has a similar mobility on SDS agarose electrophoresis to the smallest multimer of VIIIR:Ag, they are not immunologically identical.

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In Vivo and In Vitro Comparison of Various Factor VIII Concentrates

10.1055/s-0039-1682451 ◽

1977 ◽

Author(s):

I.M. Nilsson ◽

U. Hedner

Keyword(s):

Factor Viii ◽

Half Life ◽

The Other ◽

Crossed Immunoelectrophoresis ◽

Factor Viii Concentrates ◽

Fibrinogen Content ◽

In Vivo Recovery ◽

In Vitro Comparison

Five different factor VIII concentrates, AHF-Kabi(=fraction 1-0), Krynativ-Kabi(=cryoprecipitate), Hemofil-Hyland, AHF-Profilate-Abbott, Kryobulin-Immuno, available in Sweden for treatment of haemophiliacs were compared with respect to in vivo recovery of F VIII:C and survival time and in vitro properties. The parameters studied were F VIII:C, F VIIIR:AG, crossed Immunoelectrophoresis, F VIII:Rcof, fibrinogen content and F XIII activity. All the preparations had higher values for F VIIIR:AG than for F VIII:C. The quotient was highest for Hemofil, Krynativ-Kabi and Kryobulin and varied between 4 and 7. The lowest quotient, 1.3 to 4, showed AHF-Kabi. The units of F VIII:Rcof were almost the same as the units of F VIII:C. AHF-Kabi had the highest fibrinogen content and was the only preparation with high amounts of F XIII. In cross Immunoelectrophoresis AHF-Kabi showed a similar pattern to that of normal plasma. The other preparation had a different pattern suggesting less hetero-genicity of the molecule. The in vivo recovery was about the same for all the concentrates but AHF-Kabi had a significantly longer half-life (18-26 hrs); the corresponding figures for Hemofil were 8-16 hrs when given to the same patients. Only AHF-Kabi was able to completely normalize the defect in von Willebrand’s disease.

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DDAVP Factor VIII Concentrate and its Properties in VIVO and in VITRO

10.1055/s-0039-1684437 ◽

1979 ◽

Author(s):

I-M. Nilsson ◽

M. Mikaelsson ◽

H. Vilhardt ◽

B. Wiechel

Keyword(s):

Factor Viii ◽

Blood Donors ◽

In Vivo Studies ◽

Severe Haemophilia ◽

Plasma Pool ◽

Factor Viii Concentrates ◽

Qualitative Effect ◽

Fraction I

DDAVP fraction I-0 was prepared from a plasma pool derived from 80 blood donors who had received DDAVP and tranexamic acid. A control fraction I-0 was prepared from a plasma pool from the same group of blood donors under identical conditions except for the drug treatment.The DDAVP plasma pool as well as the DDAVP fraction I-0 contained 2-3 times as much VIII:C as the controls. VIIIR:Ag increased to a lesser extent. No difference in stability of VIII:C was seen. In vivo studies showed that infusion of the DDAVP fraction I-0 to 3 patients with severe haemophilia A caused a 2-3 times higher increase in VIII:C than after infusion of the same volume of the control fraction I-0. No difference in disappearance rate of VIII:C was seen.Both factor VIII concentrates normalised the defect in VWD.It is concluded that administration of DDAVP to blood donors prior to collection of blood increases the content of VIII:C in the ensuing factor VIII concentrate and secondly that such stimulation has no demonstrable qualitative effect on the factor VIII obtained.

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Specific factor VIII-related antigen fragmentation: an in vivo and in vitro phenomenon

Blood ◽

10.1182/blood.v60.4.930.bloodjournal604930 ◽

1982 ◽

Vol 60 (4) ◽

pp. 930-939 ◽

Cited By ~ 1

Author(s):

RR Montgomery ◽

J Johnson

Keyword(s):

Factor Viii ◽

Bleeding Time ◽

Antigenic Determinants ◽

Specific Factor ◽

Crossed Immunoelectrophoresis ◽

Agarose Electrophoresis ◽

Factor Viii Concentrates

Factor VIII concentrates have been shown to have reduced ability to correct the bleeding time defect in von Willebrand's disease and to have abnormal mobility of their VIIIR:Ag on crossed immunoelectrophoresis. This report concerns the partial characterization of a fragment of VIIIR:Ag that lacks some of the normal antigenic determinants present on VIIIR:Ag. It is present in commercial factor VIII concentrates, but not in cryoprecipitate, and is recovered from the plasma of hemophilic patients who have been infused with these concentrates. The fragment is also produced during disseminated intravascular coagulation. Although it has a similar mobility on SDS agarose electrophoresis to the smallest multimer of VIIIR:Ag, they are not immunologically identical.

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