This study was aimed to compare the collection efficiency of mononuclear cells (MNC) from peripheral blood as well as the changes of blood-related indices in patient by using 3 cell separators. MNC were collected from 94 tumor patients by using Fenwal CS-3000plus, Haemonetics MCSplus and COBE spectra separators. Routine blood test was performed before and after MNC collection to detect the potential effects of cell separators on blood-related indices in the patients. MNC count was performed. The percentages of CD3(+), CD4(+) and CD8(+) in peripheral blood cells were determined. The results showed that the MNC counts were (3.08 ± 0.79)×10(9), (3.21 ± 1.12)×10(9), and (3.22 ± 1.84)×10(9) per bag by CS-3000plus, MCSplus and COBE spectra, respectively. And the corresponding decrease of platelet percentage was (6.86 ± 5.70)%, (8.05 ± 5.14)% and (5.89 ± 4.48)%, respectively. The CD3, CD4 and CD8 ratios in peripheral blood of patients before and after treatment were significantly statistical different (P < 0.001). It is concluded that the MNC collection can be performed successfully with CS-3000plus, MCSplus and COBE spectra, and their collections can meet the needs in clinic.
Aged ; Cell Separation ; methods ; Cytokine-Induced Killer Cells ; cytology ; Dendritic Cells ; cytology ; Female ; Humans ; Leukapheresis ; instrumentation ; Male ; Middle Aged

OBJECTIVETo establish a new method for sperm sorting by imitating the physiological process of sperm-cervical mucus interaction on the microfluidic chip.

METHODSWe designed a microfluidic chip to imitate the physiological process of natural sperm sorting in the microchannel based on the interaction between sperm and cervical mucus, and obtained motile sperm after the interaction. Meanwhile, we established an integrated real-time sperm detection reservoir on this chip to determine sperm parameters using the computer-assisted sperm analysis system. We analyzed 30 samples using both microfluidic and swim-up methods, and compared the results with those obtained before sorting.

RESULTSThe rate of grade a + b sperm, the rate of morphologically normal sperm, straight-line velocity (VSL), average path velocity (VAP) and straightness (STR) were (29.78 +/- 11.24)%, (8.00 +/- 5.19)%, (18.89 +/- 4.90) microm/s, (26.84 +/- 5.13) microm/s and (70.15 +/- 7.61)%, respectively, before sorting, (71.65 +/- 11.18)%, (14.95 +/- 6.79)%, (24.14 +/- 5.95) microm/s, (32.61 +/- 6.36) microm/s and (73.87 +/- 9.34)%, respectively, after swim-up sorting, and (92.37 +/- 6.33)%, (23.33 +/- 7.67)%, (34.03 +/- 16.78) microm/s, (38.73 +/- 16.40) microm/s and (84.91 +/- 12.56)%, respectively, after sorting on the microfluidic chip. The sperm parameters obtained before sorting showed statistically significant differences from those obtained on the chip (P < 0.01) and by the swim-up method (P < 0.05).

CONCLUSIONImitation of the physiological interaction between sperm and cervical mucus on the microfluidic chip helped the realization of both the natural sorting and real-time analysis of sperm. The quality of the sperm sorted on the microfluidic chip is significantly better than that of the sperm before sorting and sorted by the swim-up method. This has prepared the ground for imitating the fertilization process under the physiological condition on the microfluidic chip.

Cell Movement ; Cell Separation ; Cervix Mucus ; Humans ; Male ; Microfluidic Analytical Techniques ; instrumentation ; Microfluidics ; methods ; Oligonucleotide Array Sequence Analysis ; Semen Analysis ; Sperm Motility ; physiology ; Spermatozoa ; physiology

OBJECTIVETo investigate the effects of a microfluidic sperm sorter on the routine parameters and DNA integrity of human sperm.

METHODSWe divided 40 semen samples into two aliquots and performed sperm sorting using a self-made polydimethylsiloxane microfluidic sperm sorter and the swim-up method, respectively. Then we evaluated and compared the effects of these two methods on the sperm routine parameters and DNA integrity by computer-assisted sperm analysis and sperm chromatin dispersion test.

RESULTSAfter processing, sperm motility, normal morphology and tail hypoosmotic swelling rate were significantly improved, while sperm DNA damage remarkably decreased (P < 0.01). The microfluidic sperm sorter achieved a significantly lower rate of sperm DNA damage than the swim-up method ([ 8.4 +/- 5.8 ]% vs [16.4 +/- 9.2] %, P < 0.01), but no statistically significant differences were found in all other parameters between the two methods.

CONCLUSIONHigh-quality sperm with less DNA integrity damage could be obtained in sperm sorting with the microfluidic sperm sorter.

Adult ; Cell Separation ; instrumentation ; methods ; DNA ; DNA Damage ; Humans ; Infertility, Male ; genetics ; physiopathology ; Male ; Microfluidics ; Middle Aged ; Protein Array Analysis ; Semen Analysis ; instrumentation ; methods ; Sperm Motility ; Spermatozoa

BACKGROUND: The bone marrow (BM) cellularity has been used as a major index for the evaluation of hematologic diseases and malignancies. However, the microscopic evaluation lacks objectivity because of considerable variations among different observers. We measured myelocrit cellularity as an objective method to evaluate the cellularity. METHODS: Between November 2007 and January 2008, 489 consecutive BM aspirates including 25 cases of AML D7 marrow (7 days after initiation of chemotherapy) were examined. The conventional BM cellularity was evaluated using BM needle biopsies after hematoxylin-eosin stain. EDTA-anticoagulated BM aspirates were put into the Wintrobe tubes, centrifuged and the thickness of 5 layers from the bottom was measured: RBC, buffy coat (BC), plasma, BM particle, and fat layers. The myelocrit cellularity was defined as the ratio of BC to the BC plus fat layers. RESULTS: Both of the thickness of BC layer (r=0.721, P=0.000) and the myelocrit cellularity (r=0.735, P=0.000) correlated well with the conventional cellularity. However, the AML D7 BM cellularity correlated with BC layer (r=0.589, P=0.002), but not with the myelocrit cellularity (r=0.281, P=0.231). The cellularity of the BM other than AML D7 marrow showed a better correlation with the myelocrit cellularity (r=0.826, P=0.000) than the BC layer (r=0.713, P=0.000). CONCLUSIONS: The myelocrit is a simple, reproducible and objective method to determine the BM cellularity. For accurate assessment of BM cellularity, measurement of the thickness of BC layer in AML D7 BM and of the myelocrit cellularity in other BM samples has better be used.
Adolescent ; Adult ; Aged ; Bone Marrow/*pathology ; Cell Separation/instrumentation/*methods ; Child ; Child, Preschool ; Female ; Humans ; Infant ; Leukemia, Myeloid, Acute/*pathology ; Male ; Middle Aged

OBJECTIVETo establish an semi-automated effective method for large-scale purification of islet cells from human pancreas.

METHODSHuman pancreas tissue was digested with collagenase P using a semi-automated pancreas-digestion system followed by purification in a HCA-Ficoll continuous gradient using Cobe2991 cell separator. After isolation, the islet cell yield and purity was evaluated with light microscope with DTZ staining, and the islet function assessed by insulin release assay in vitro.

RESULTSThe number of the islets collected from each pancreas averaged 38 6201-/+78 219 islet equivalents (IEQ) before purification, and 231 420-/+28 054 IEQ after the purification with discontinuous gradient centrifugation. From each gram of the pancreatic tissue, 3148-/+317 IEQ were obtained with an average purity of (62.81-/+2.68) %. The purified islets responded well to high-concentration (16.7 mmol/L) glucose stimulation with a 2.53-fold increase of insulin secretion over the basal level (3.3 mmol/l, P<0.001).

CONCLUSIONThe established semi-automated method can be applicable for large-scale purification of fully functional islet cells from human pancreas.

Cell Count ; Cell Separation ; instrumentation ; methods ; Cell Survival ; Glucose ; pharmacology ; Humans ; Insulin ; secretion ; Islets of Langerhans ; cytology ; drug effects ; metabolism ; Reproducibility of Results

This study was aimed to compare the difference of quality between buffy-coat-collected platelet concentrates (BC-PC) and single-donor plateletpheresis (SDP). 15 packs of BC-PC and 15 units SDP were stored at 20 degrees C - 24 degrees C with agitation. Platelet concentration, platelet volume, residual leukocyte and residual erythrocyte in two groups were examined after preparation for 1 hour. Mean platelet volume, pH value, hypotonic shock response (HSR), CD62p expression and CD62p re-expression of platelet were detected on 0, 1, 2, 3, 4, 5 days of platelet preservation. The results showed that the platelet yields, residual leukocyte and residual erythrocyte in two groups accorded with the national quality standard respectively, but residual leukocyte and residual erythrocyte in BC-PC group were higher than those in SDP group when platelet yields in two groups were equal (p < 0.01). Lactate concentration, CD62p expression of platelet increased with prolongation of preseved time, while pH value decreased gradually. Compared with SDP group, there were significant differences in CD62p expression, CD62p re-expression of platelet preserved for 0 - 5 days (p < 0.01), and in pH value of platelet preserved 2 - 5 days (p < 0.01). There was no changes in HSR of SDP group for 0 - 5 days, while HSR in BC-PC group decreased gradually. There were significant differences in HSR of platelet preserved for 1 - 5 days (p < 0.01). It is concluded that the platelet concentrates prepared by BC-PC are not equal to SDP in quality, the preparation technology of BC-PC should be optimized further in order to reduce residual leukocyte, residual erythrocyte and activated platelet yields, as well as improve the quality of BC-PC.
Blood Platelets ; metabolism ; physiology ; Blood Preservation ; Cell Separation ; methods ; Erythrocytes ; cytology ; Humans ; Lactic Acid ; blood ; Leukocytes ; cytology ; P-Selectin ; blood ; Platelet Count ; Plateletpheresis ; instrumentation ; methods ; Quality Control

To evaluate the yield of the blood cell separator for collection of peripheral blood stem cells (PBSC) from ABO major and (or) minor incompatible allogeneic donors and the feasibility of PBSC component infusion to the recipients without removal of erythrocytes or plasma, the Cobe Spectra (Version 6.1) blood cell separator was utilized to collect PBSC component from 9 allogeneic donors. Of all the donors, 4 were ABO major incompatible, 2 were minor incompatible and the other 3 were both major and minor incompatible to corresponding recipients. In each cycle, different amount of PBSC component was harvested, and the variable volume plasma chased the cells into the bag was adjusted according to the ABO incompatibility. The nucleated cell count, percentage of mononuclear cell, number of CD34(+) cell and percentage of viable cell (trypan blue excluding rate) in the component were detected. At the time of infusion, a series of protective measures to the renal function of recipients were taken. The results showed that apheresis was twice performed on these eight donors to collected enough PBSC for transplantation or cryopreservation, except one apheresis was enough for cell amount needed by transplantation, as the donor's body weight was much heavier than that of the recipient. Altogether 17 apheresises were performed, the mean yield of nucleated cells was 3.77 x 10(10), in which 97% to 99% were mononuclear cells (MNC). The harvested number of CD34(+) cell was 8.62 x 10(6)/kg. All the trypan blue exclusion rate was 100%. In ABO major incompatible or both major and minor incompatible component, there were 8 - 10 ml packed erythrocytes; in ABO minor incompatible component, there were 80 - 120 ml of plasma. These components were infused into the recipients without removal of erythrocytes or plasma and no haemolytic reaction was observed in any recipient, and their hematopoietic functions soon recovered. Results suggest that enough PBSC can be acquired by using blood cell separator Cobe Spectra (Version 6.1), with the modified separation factors, and the collected PBSC component can be safely infused into the ABO incompatible recipients without removal of erythrocytes or plasma.
ABO Blood-Group System ; Antigens, CD34 ; blood ; Blood Component Removal ; methods ; Blood Donors ; Blood Group Incompatibility ; blood ; Cell Separation ; instrumentation ; Cell Survival ; Hematopoietic Stem Cell Mobilization ; Hematopoietic Stem Cells ; cytology ; immunology ; Humans ; Peripheral Blood Stem Cell Transplantation ; methods ; Transplantation, Homologous

To evaluate the hematopoietic stem/progenitor cell apheresis effect of Cobe Spectra (Version 6.1) and Fenwal CS 3000 Plus cell separators, fourty-two procedures on twenty donors using Cobe Spectra cell separator and twenty-two procedures on sixteen donors using Fenwal CS 3000 Plus cell separator were retrospectively analyzed. The number of CD34(+) cells collected, the collection efficiency (CE) of CD34(+) cells and the contaminations of red blood cell and platelet in the stem/progenitor cell products of two devices were compared. The results showed that there were no significant differences in the total number of CD34(+) cells collected and the CD34(+) cell CE between the two devices. There were positive correlations between the count of peripheral blood cells including leukocyte, monocyte, hematopoietic progenitor cell and CD34(+) cell after mobilization and the total number of CD34(+) cells collected. The stepwise multiple variable analyses revealed the peripheral blood stem/progenitor cell count emerged as the only significant independent predictive factor for CE. A negative correlation was seen between the peripheral blood monocyte count and the CD34(+) cell CE for the Fenwal CS 3000 Plus. The Fenwal CS 3000 Plus product contained more red blood cells than that of the Cobe Spectra. The decrease in the peripheral platelet count after Fenwal CS 3000 Plus apheresis was also greater. It is concluded that collection efficacy of Cobe Spectra (Version 6.1) and Fenwal CS 3000 Plus was similar. Cobe Spectra shall be used preferably to assure higher CD34(+) cell CE at a high peripheral blood monocyte count. The Cobe Spectra cell separator is better for the donors with mismatched blood type and the donors with thrombocytopenia.
Antigens, CD34 ; blood ; Blood Cell Count ; Cell Separation ; instrumentation ; methods ; Hematopoietic Stem Cell Mobilization ; Hematopoietic Stem Cells ; cytology ; Humans ; Leukapheresis ; instrumentation ; Reproducibility of Results ; Retrospective Studies

A protocol for enrichment and adsorption of karyocyte from whole blood by using magnetic nanometer beads as solid-phase absorbents was presented. The PCR amplification could be accomplished by using the nanobeads with karyocyte as template directly and the PCR products were applied on an oligonucleotide array to do gene typing. The HLA-A PCR amplification system and a small HLA-A oligonucleotide microarray were applied as the platform and an experiment protocol of separating karyocyte from whole blood using the magnetic nanometer beads (Fe2O3) were set up. The experimental conditions were also discussed. It showed that pH level of PBS eluent, Taq enzyme quantity and fragment length of products could influent the amplification results, and the magnetic nano-beads could succeed in sample preparation in microarray to provide a promising way in automatic detection and lab-on-a-chip.
Adsorption ; Cell Separation ; instrumentation ; methods ; Genotype ; Leukocytes ; cytology ; Magnetics ; Microchemistry ; instrumentation ; methods ; Nanotechnology ; Oligonucleotide Array Sequence Analysis ; methods ; Polymerase Chain Reaction ; Templates, Genetic

A protocol for enrichment and adsorption of karyocyte from whole blood by using magnetic nanometer beads as solid-phase absorbents was presented. The PCR amplification could be accomplished by using the nanobeads with karyocyte as template directly and the PCR products were applied on an oligonucleotide array to do gene typing. The HLA-A PCR amplification system and a small HLA-A oligonucleotide microarray were applied as the platform and an experiment protocol of separating karyocyte from whole blood using the magnetic nanometer beads (Fe2O3) were set up. The experimental conditions were also discussed. It showed that pH level of PBS eluent, Taq enzyme quantity and fragment length of products could influent the amplification results, and the magnetic nano-beads could succeed in sample preparation in microarray to provide a promising way in automatic detection and lab-on-a-chip.
Adsorption ; Cell Separation/instrumentation ; Cell Separation/methods ; Genotype ; Leukocytes/*cytology ; *Magnetics ; Microchemistry/instrumentation ; Microchemistry/*methods ; Nanotechnology ; Oligonucleotide Array Sequence Analysis/*methods ; Polymerase Chain Reaction ; Templates, Genetic