Abstract:Metaproteomics is an emerging proteomics technology to analyze large scale protein expression in environmental microbial ecosystem. It is termed as the large-scale characterization of the entire protein complement of environmental microbial community at a given point in time. This review focuses on the research strategies and the recent applications in this field based on the published reports and in combination with our own research experiences.

Abstract:To establish human β-defensin-3 gene transgenic cell lines as competent donor cells for the production of transgenic animals using somatic cell nuclear transfer (SCNT). Firstly, we obtained human β-defensin-3 by RT-PCR from human placenta, and subsequently inserted the fragment hBD into the corresponding site of the plasmid pBCP. Then we moved the combined fragment BCD (including 5￠ and 3￠ regulating region of β-casein and hBD) into the corresponding site of the plasmid pEGFP-C1. Finally we successfully constructed mammary-specific expression vector pEBCD. We transected pEBCD into Holstein Fetal fibroblast cells by Lipofectamine TM-2000 and selected in medium with G418 for three to four weeks. We identified G418 resistant transfectants by PCR, RT-PCR and EGFP detection. Our results indicated that human β-defensin-3 gene stably was integrated into the open region of the chromatin in G418 resistant fibroblast cells. Meanwhile we identified the expression of human β-defensin-3 in the supernant of stable transfected mammary epithelial cells by Western blotting. This study may provide competent transgenic donor cells for the production of transgenic animals by SCNT and improve the efficiency of transgenic cloning.

Abstract:In order to get new antibacterial peptide, we designed a hybrid peptide LfcinB(1-15)-Melittin(5-12), composed of 1-15 amino acid residues of bovine Lactoferricin and 5-12 amino acid residues of Melittin. According to the bias of codon utilization of Escherichia coli, We synthesized the gene encoding the hybrid peptide. We inserted the gene between the sites of Nco ? and Sal ? of pET-32a and obtained the recombinant expression vector for heterologous expression of LfcinB(1-15)-Melittin(5-12) in Escherichia coli. We used Escherichia coli BL21(DE3) as expression host for the recombinant plasmid. After induced by isopropyl-β-D-thiogalactoside (IPTG) under the optimized conditions, we realized the fusion protein was successfully expressed. The fusion protein was expressed in soluble form and the level was more than 35% of the total proteins. With (His)6×Tag, the fusion protein was easily purified by His×Bind Purification Kit. After purification, we obtained 35 mg of fusion protein from 1 L of culture medium. At last, we accomplished that the peptide LfcinB(1-15)-Melittin(5-12) was released from the fusion protein cleaved by enterokinase. The recombinant LfcinB(1-15)-Melittin(5-12) showed antimicrobial activity assayed by agar diffusion test. This is the first report on the heterologous expression of the hybrid antibacterial peptide LfcinB(1-15)-Melittin(5-12) in Escherichia coli and also provides basis for next cost-effective expression of other antimicrobial peptides in genetic engineering.

Abstract:After sequencing, we amplified and cloned foot-and-mouth disease virus (FMDV) O/QYYS/s/06 whole genome by three fragments. These three fragments were cloned into vector P43 one by one to construct recombinant plasmid P43C, which carried the full-length cDNA of FMDV O/QYYS/s/06. Then, plasmid P43C and plasmid T7 expressing T7 RNA polymerase were co-transfected into BHK-21 cells. After 48 h, we harvested the culture broth from transfected BHK-21 cells and inoculated into 2-3 day-old sucking mice. After four generation passage, the virus harvested from sucking mice was confirmed to be type O FMDV by the indirect hemagglutination test, sucking mice’s neutralization test and sequencing. The results showed that we have successfully constructed the full-length cDNA clone of FMDV O/QYYS/s/06 strain.

Abstract:We studied the immunogenicity of pseudorabies virus gC DNA vaccination by fusing the murine complement C3d receptor binding domain. First, pseudorabies virus gC gene was linked to four copies of C3d receptor binding domain (M284), and then cloned into the vector pcDNA3.1 to construct the recombinant plasmid sgC-M284. Through the experiment of immunized BALB/c mice, we found that the enzyme linked immunosorbent assay (ELISA) antibody titer for sgC-M284 was 17-fold higher than that for sgC alone, and protective rate of mice was augmented from 25% to 88% after lethal dose PrV (316 LD50) challenge. In addition, the IL-4 levels for sgC-M284 immunization approached that for the pseudorabies virus inactivated vaccine. In conclusion, we demonstrated murine C3d receptor binding domain fusion significantly increased Th2-biased immune response by inducing IL-4 production.

Abstract:We evaluated the effect of biological pretreatment with white rot fungus Trametes vesicolor on the enzymatic hydrolysis of two wood species, Chinese willow (Salix babylonica, hardwood) and China-fir (Cunninghamia lanceolata, softwood). The result indicated that the pretreated woods showed significant increases in the final conversion ratios of enzymatic hydrolysis (4.78-fold for hardwood and 4.02-fold for softwood). In order to understand the role of biological pretreatment we investigated the enzyme-substrate interactions. Biological pretreatment enhanced the substrate accessibility to cellulase but not always correlated with the initial conversion rate. However, the change of the conversion rate decreased dramatically with increased desorption values after biological pretreatment. Thus, the biological pretreatment slowed down the declines in conversion rates during enzymatic hydrolysis by reducing the irreversible adsorption of cellulase and then improved the enzymatic hydrolysis. Moreover, the decreases of the irreversible adsorption may be attributed to the partial lignin degradation and alteration in lignin structure after biological pretreatment.

Abstract:To study the role of the thyroid hormone receptor TRaA involved in the process of the metamorphic development of Japanese flounder, we firstly cloned the TRαA gene, then ligated into the fusion expression vector pET30a and expressed in Escherichia coli DE3 (BL21) host cells. After induced for 4 h with 1 mmol/L Isopropyl b-D-Thiogalactoside, the target fusion protein was successfully expressed and identified in inclusion bodies by SDS-PAGE and Western blotting. The recombinant protein was denatured and purified by His-Bind resin, then renatured through gradient washing on His-bind resin column. After that, polyclonal antibody was prepared by immunizing New Zealand rabbits with purified protein. Dot blotting analysis showed the antibody with the titer of 1:200 000 reacted specifically to the expressed recombinant protein. Furthermore, a chromatin immunoprecipitation assay was performed to identify the specific binding between the antibody and TRαA in living cells of Japanese flounder. The result showed that thyroid hormone was involved in the alkaline phosphatase (ALP) gene transcriptional regulation through TRaA in vivo.

Abstract:Acetohydroxyacid synthase (AHAS) catalyses the first reaction in the pathway for synthesis of the branched-chain amino acids. AHAS is the target for sulfonylurea, imidazolinone and other AHAS-inhibitor herbicides. Herbicides-resistant AHAS genes have potential application in plant transgenetic engineering and development of new generation herbicide. The AHAS isozyme genes ilvBN, ilvGM and ilvIH were cloned from metsulfuron-methyl resistant strain Klebsiella sp. HR11 and metsulfuron-methyl sensitive strain Klebsiella pneumoniae MGH 78578. Homologous sequences comparison indicated that the differences in AHAS isozyme genes at amino acid levels between strain HR11 and strain MGH 78578 were mainly on the large subunits of ilvBN and ilvGM. The three AHAS isozyme genes from HR11 and MGH 78578 were ligated into the expression vector pET29a(+) and expressed in Escherichia coli BL21, respectively. The results of enzyme inhibition assay showed that only ilvBN and ilvGM from strain HR11 showed strong resistance to AHAS-inhibitor herbicides, while ilvIH from strain HR11 and ilvBN, ilvGM and ilvIH from strain MGH 78578 were sensitive to AHAS-inhibitor herbicides.

Abstract:Arylalkylamine N-acetyltransferase (AANAT) and Hydroxyindole O-methyltransferase(HIOMT) are the key regulation enzymes in the melatonin biosynthesis pathway in mammals. The AANAT and HIOMT genes were constructed into a binary plant expression vector YXu55. Using leaf strips as the recipiences, we efficiently transformed tobacco (Nicotiana tabacum) variety qinyan 95 by the Agrobacterium mediated method. After gradient selection with gentamycin, a number of transgenic plants were regenerated. Southern blot and RT-PCR analyses showed that the AANAT-HIOMT genes were integrated into the genome of the transgenic plants and the target genes could express at the level of RNA transcription. By RP-HPLC, we measured the melatonin contents in transgenic plants. The results showed that the melatonin level in YXu55 (containing the gentamycin-resistance gene, the AANAT gene and HIOMT gene) transgenic plants were much higher than those in pZP122 (control containing only the gentamycin-resistance gene) transgenic plants and nontransgenic plants. The content of melatonin in pZP122 transgenic plants was nearly the same as that in nontransgenic plants. Physiological determination of antioxidative characteristics demonstrated that 1) the capacity of total antioxidation, 2) the activities of superoxide dismutase (SOD), peroxidase (POD) and catalase (CAT) and 3) the content of glutathione (GSH) were increased in YXu55 transgenic plants containing the AANAT-HIOMT genes as compared to the control plants (pZP122 or nontransgenic plants). At the same time, malonaldehyde (MDA) content did not appear remarkably difference between transgenic plants and nontransgenic plants. The above mentioned facts indicate enhancement of melatonin levels in YXu55 transgenic plants might help to reduce damage by oxidative stress.

Abstract:Prion leads to fatal transmissible spongiform encephalopathies. Cellular prion protein (PrPC) is necessary in prion disease. At present, it is demonstrated that PrPC plays a protective role in several carcinomas, such as gastric and breast cancer. We designed four 19-nt siRNAs according to cDNA sequence of human PrPC and constructed retrovirus-based RNAi vectors. We evaluated the inhibitive effect of these sequences on HuPrPC (human PrPC) and selected out three sequences with stable and efficient inhibition. And the efficiency of si626 reached more than 85%, which effect was significant. Next, we performed cell invasion assays of PC3M-si292 and PC3M-si626 in which PrPC was inhibited. And it showed that the cell invasive ability decreased in PrPC knock-down cell lines. This will make preparations for the further research on gene therapy of prion diseases and PrPC related carcinoma treatment and PrPC could be considered as a potential therapeutic target molecule in prostate cancer treatment.

Abstract:Rhesus macaque (Macaca mulatta) is the best model to study of human immunodeficiency virus (HIV) infection and to develop acquired immunodeficiency syndrome (AIDS) vaccine. The crystal structure of its major histocompatibility antigen complex (MHC) is helpful to understand the mechanism of HIV immune evasion. In this study, we cloned the light chain (β2m) of MHC class I allele of rhesus macaques, Mamu-A*02, and inserted it into pET21a(+) vector. We transfected the recombinant plasmid pET21a(+)-Mamu-β2m and pET21a(+)-Mamu-α into BL21(DE3). Mamu-A*02 and β2m were expressed in the form of inclusion bodies in BL21 (DE3). We co-refolded the inclusion bodies of Mamu-α and Mamu-β2m with SIV nonapeptide YY9 and obtained the correct refolded protein complex. Then we purified the protein complex by the gel filtration and anion-exchange column. With hanging-drop method, we screened and optimized for the protein crystal. We managed to collect a X-ray diffraction with the resolution to 2.8 ? in the condition of 0.1 mol/L BIS-TRIS (pH5.5), 2.0 mol/L(NH4)2SO4. This crystal belong to perpendicular space group P212121, with unit-cell parameters a=128.99 ?, b=129.01 ?, c=129.03 ?. This data is available for the structure determination.

Abstract:We constructed the recombinant adenovirus vector expressing IL-24 and E1A (Ad-IL-24-E1A) and investigated the inhibition of Ad-IL-24-E1A on SMMC-7721 hepatocellular carcinoma in vitro. We amplified IL-24 gene by PCR using pAdTrack-IL-24 as template. The IL-24 gene was cloned into pAdTrack-IRES at the BglⅡand SalⅠsite to form pAdTrack-IL-24-IRES. E1A digested from pAdTrack-E1A was cloned into the pAdTrack-IL-24-IRES at the Xho I and EcoR V site to form the pAdTrack-IL-24-IRES-E1A. We co-transformed both pAdTrack-IL-24-IRES-E1A and pAdeasy-1 digested by Pme I and packaged to obtain Ad-IL-24-E1A. Ad-IL-24-E1A at 50 MOI infected SMMC-7721 cells. 3-(4,5-Dimethylthiazol-2-yl)-2,5- diphenyltetrazolium bromide (MTT) assay determined cell proliferation. Flow cytometry detected Cell apoptosis. The apoptotic rate of SMMC-7721 cells was 52% 48 h after infection with Ad-IL-24-E1A. The result showed that the growth of SMMC-7721 cells was significantly inhibited by Ad-IL-24-E1A at the MOI of 50.

Abstract:We constructed and expressed an anti-CD3/anti-Pgp (P-glycoprotein) diabody previously. However, the two chains of diabody are associated non-covalently, resulting in being capable of dissociating. The aim of this study is to enhance the stability of the diabody. We introduced cysteine residues into the CD3 or Pgp V-domain to covalently lock the two chains together. The disulphide crosslinked diabody were expressed by Escherichia coli (E. coli) 16C9 and purified by a cation exchange column and an anti-Etag affinity chromatography. The purified proteins were verified through SDS-PAGE. Flow cytometry (FCM) was used to analyse the binding properties, competitive binding capacity and stability in vitro. The dsPpg-diabody failed to form disulphide bond properly. The designed disulphide bridge between the different chains of dsCD3-diabody was formed correctly. FCM demonstrated the dsCD3-diabody has specific antigen binding activity, the same binding activity and competitive binding activity as its parent diabody. The dsCD3-diabody retained the full activity even after 72 h incubation at 37°C in human serum, in contrast, the parent diabody began to lose activity after only 1 h and lose all its activity 24 hours later. The induced disulphide bond in the CD3 V-domain effectively enhanced the stability of anti-CD3/anti-Pgp diabody. The method of stabilizing a diabody by introducing a disulphide bond into is practical.

Abstract:PRAS40 (proline-rich Akt substrate 40 kD) associates with mammalian target of rapamycin complex 1(mTORC1), serine 183 site (Ser183) of PRAS40 can be phosphorylated by mTORC1. To prepare the phosphorylated PRAS40 (Ser183) antibody, We chosen 10-amino acid including Ser183 as antigen peptide through antigenicity and hydrophobicity analysis, hinged on keyhole limpet hemocyanin (KLH), and used the KLH-peptide to immunize rabbits. After antibody serum titer detection by enzyme linked immunosorbent assay (ELISA), the antibody was purified with rProtein A sepharose fast flow and dephosphorylated antigen membrane. The antibody titrate reached 1:10 000 after purification and its special property was enhanced with absorption treatment of dephosphorylated antigen membrane. In addition, we used rabbit anti-PRAS40 antibody and the phosphorylated PRAS40 (Ser183) antibody to detect PRAS40 expression in several cell lines, including the normal cells HL7702, HEK293, tumor cells HepG2, A549 and S180. There were no quite difference among these cells; otherwise, we observed the decreased phosphorylation level of Ser183 after amino acid withdrawal treatment. Therefore, the polyclonal phosphorylated PRAS40 (Ser183) antibody was specific to PRAS40 (Ser183) site and could be used for the function study of PRAS40.

Abstract:We immobilized carbonic anhydrase (CA) onto the surface of membrane oxygenator of polymethyl pentene (PMP) to enhance the removal of carbon dioxide in blood by two steps. We first introduced hydroxyl groups onto PMP surface by water plasma treatment, and then coupled CA onto PMP surface by using cyanate bromide (CNBr) as a crosslinker. After plasma treatment, the contact angle with water and chemical composition of PMP surface were characterized by analysis system of surface contact angle and XPS. Using p-nitrophenyl acetate (p-NPA) as a substrate, the activity, concentration, storage stability and re-usability of immobilized CA on PMP hollow fibers were studied by ultraviolet spectrophotometer. The preliminary data showed that hydroxyl groups could be introduced on the surface of PMP by water plasma treatment, and CA with catalysis activity could be successfully introduced onto PMP surface in high immobilization efficiency. The activity of covalently immobilized CA increased with the increase of concentration of CNBr, and the maximum was 73% of the theoretical activity of CA spread on PMP surface in monolayer in studied range. Covalently immobilized CA showed higher reusability compared to physically adsorbed CA, and higher storage stability compared to free CA in solution at 37oC. The method would be used potentially in the membrane oxygenator to improve the capacity of removal of carbon dioxide in blood in the future.

Abstract:Polysaccharides from Dendrobium huoshanense possess immunostimulating activity, antioxidant activity and anticataract activity. In order to produce the active polysaccharides from Dendrobium huoshanense through cell culture, we investigated the effects of salicylic acid on cell growth, accumulation of polysaccharides and utilization of carbon source in suspension cultures of protocorm-like bodies from Dendrobium huoshanense. Although salicylic acid slightly inhibited the cell growth, it was beneficial to the utilization of carbon source, thus leading to significant increase in the contents of polysaccharides. The highest polysaccharide production occurred on the medium supplied with 100 μmol/L salicylic acid. After 18 days of culture the production of polysaccharides reached 3.129 g/L, which was 1.63 times that of the control. Further, we established the kinetic models describing cell growth, polysaccharide production and carbon source utilization based on Logistic equation, Luedeking-Piret equation and Luedeking-Piret-Like equation. The calculated values from the kinetic models showed a good fit to the experimental values, suggesting that salicylic acid could be an effective compound to enhance the production of active polysaccharides from protcorm-like bodies from Dendrobium huoshanense.

Abstract:As an immunodepressant, anti-CD25 monoclonal antibody has a huge market with wide prospect and economic value. We developed a low protein serum-free medium for large-scale GS-NS0 myeloma cell culture and anti-CD25 monoclonal antibody production. Further study focused on the characteristics of GS-NS0 cell growth, glucose and amino acid metabolism, and antibody production. In the serum-free medium, the maximal viable cell density and antibody concentration reached above 3×106 cells/mL and 300 mg/L in batch culture. Compared with the commercial serum-free medium (Excell 620 + 0.2% Primatone), the maximal viable cell density doubled and the maximal antibody concentration increased 46%. Results also showed the specific growth rate decreased when the glucose concentration was lower than 6 mmol/L. And the production of lactate increased when glucose concentration was excessively high (>30 mmol/L). These results were important to provide technique and theory basis for developing optimized GS-NS0 cell culture and anti-CD25 monoclonal antibody production processes.

Abstract:Laser scanning confocal microscope (LSCM) is currently the only equipment to observe fluorescence. However, this technique has disadvantages such as high cost and long test process. In this study, we developed a new system of laser-induced fluorescence (LIF) for microfluidic chip applied to detecting the expression of green fluorescent protein (GFP) in Bacillus subtilis. This novel system was comprised of laser device, optics unit, microfluidic chip, photomultiplier and computer treatment unit. The tests indicated that microfluidic chip could detect the expression of GFP as sensitively as LSCM in Bacillus subtilis. Moreover, this LIF detection system could instead of PCR to identify the positive clone in this special case. Nevertheless, the LIF system only was suitable to detect the fluorescent strength of GFP, and could not meet the request of some cases for example protein location. Therefore, this system will be applied in environmental detection with microbe, drug discovery and other cases.

Abstract:Here, we presented a method to bacterially express the major structural protein L1 of Human Papillomavirus type 18 (HPV18) as soluble form. We found that the purified L1 could self-assemble to virus-like particles (VLPs). Further, we investigated the immunogenicity and the induced level of neutralizing antibody using these VLPs. First, the genome of HPV18 was cloned from a patient in Xiamen. It was used as template for PCR amplification of HPV18 L1 gene. The resultant DNA fragment was inserted into expression vector pTrxFus and expressed in Esoherichia coli GI724. Second, L1 protein was purified by ammonium sulfate precipitation, ion-exchange chromatography and hydrophobic interaction chromatography; and the purified L1 was subjected to self-assembly to form VLPs with the removal of premixed reductant DTT. Finally, the size and morphology of these VLPs was investigated by Dynamic Light Scattering and Transmission Electronic Microscopy as 29.34 nm in hydrated radius and globular particles similar with native HPV18. The half effective dosage (ED50) and maximum level of neutralizing antibody elicitation were measured by vaccinations on mice, rabbit and goat using pseudovirus neutralization cell model. The results showed that the ED50 of HPV18 VLPs is 0.006 μg in mice, and the maximum titer of neutralizing antibody elicited in rabbit and goat is up to 107. As a conclusion, we can provide HPV18 VLPs with highly immunogenicity from prokaryote expression system, which may pave a new way for research and development of prophylactic vaccine for HPV18.

Abstract:Estrogen Receptor (ERα) is a member of superfamily of ligand-activated transcription factors which play critical roles in many biological processes. To screen novel modulators of ERα for drug development and biological function research, we developed a mammalian one-hybrid-based high-throughput screening model for ERα modulator. We cloned the ERα LBD gene from the total mRNA of fat tissue by RT-PCR and fused it with the GAL4 DNA binding domain of pBIND-GAL4 plasmid to construct a chimara expression plasmid pBIND-GAL4-Erα(LBD). The L02 cells was cotransfected with pBIND-GAL4-ERα(LBD) and a GAL4-responsive luciferase reporter plasmid pGL3-GAL4, and following treatment with test compounds for 24 h, the activities of luciferase were detected to evaluate the transactivities of ERα modulators. After manner optimizations of transfection conditions, Estradiol, an agonist control, induced the expression of luciferase in a dose-dependent with EC50 of 0.17 mmol/L, the maximum folds of induction was about 28.1. Tamoxifen, an antagonist control, efficiently suppressed the estradiol-mediated luciferase induction with EC50 of 0.10 mmol/L. Using this screening model, we discovered four ERα agonists from 2000 natural and synthetic compounds.

Abstract:The purpose was to optimize the vitrification for porcine embryos cryopreservation. Blastocyst/Morula(5-6th day-embryos) were collected from superovulated Bama mini-pigs(sows/gilts). We compared different cryopreservation methods, cryopreservation tools, thining of zona pellucida(ZP) and recipient breeds on the efficiency of porcine embryo cryopreservation. The results showed that: in embryo survival rate and blastocyst cell number, there were no significant differences between cryopreservation method I [embryos were vitrified by two step method with open pulled straw (OPS) and glass micropipette (GMP) in solution 1(TCM199 + 20% FBS + 10% EG + 10% DMSO) for 3 min, and solution 2(TCM199 + 20% FBS + 20% EG + 20% DMSO + 0.4 mol/L SUC)for 1 min, stored in liquid nitrogen] and method II[Blastocysts were cultured for 25 min in NCSU23 + 7.5 mg/mL cytochalasin B, centrifuged at approximately 13 000 ×g for 12-13 min, and recovered back into pNCSU23. They were then equilibrated for 5 min in 2 mol/L ethylene glycol in pNCSU23, washed quickly in the vitrification medium, 8 mol/L ethylene glycol, 7% polyvinylpyrrolidone (PVP) in pNCSU23, loaded into OPS/GMP, and plunged into liquid nitrogen). GMP vitrification method was more suitable and efficient than OPS method (P<0.05) in embryo survival rate (83.8% vs 77.6%) and blastocyst cell number (53.1 vs 47.5) after thawing. Thining of ZP did not increase the survival rate, but significantly improved blastocyst cell number in the survival blastcysts (60.1 and 46, P<0.01). Local pig breeds (Fengjing sows) were more suitable as recipients for embryo transfer of vitrified/warmed blastcysts, which can improve pregnant rate and embryo efficiency.

Abstract:We studied the ligation of coagulation factor VIII heavy and light chain in Escherichia coli by utilizing the intein-mediated protein trans-splicing. A B-domain deleted factor VIII (BDD-FVIII) gene was broken into two halves of heavy and light chains before Ser1657 which meets the splicing required conserved residue and then fused to 106 and 48 amino acid-containing N-part termed Int-N and C-part termed Int-C coding sequences of split mini Ssp DnaB intein respectively. These two fusion genes were constructed into a prokaryotic expression vector pBV220. Through induction for expression of recombinant protein it displayed an obvious protein band as predicted size of BDD-FVIII protein on SDS-PAGE gel. Western blotting using factor VIII specific antibodies confirmed that this protein band is BDD-FVIII produced by protein trans-splicing. It demonstrated that the heavy and light chains of BDD-FVIII can be efficiently ligated with the Ssp DnaB intein-mediated protein trans-splicing. These results provided evidence for encouraging our ongoing investigation with intein as a means in dual AAV vectors carrying the factor VIII gene to overcome the packaging size limitation of a single AAV vector in hemophilia A gene therapy.

Abstract:Alternations of lymphocyte in biophysical properties (e.g., morphology and viscoelasticity) are related to the human health, disease diagnosis and treatment. Here, we used atomic force microscopy (AFM) to characterize the morphology and mechanical properties of normal lymphocyte and Jurkat. The AFM images revealed that their cell shapes appeared similar. The mechanical properties of the two groups were tracked with AFM-based force spectroscopy. The normal lymphocyte cells had a high adhesion force distribution in (796.7±248.5) pN, whereas the Jurkat cells had a low force distribution in (158.5±37.5) pN. The adhesion force revealed that the Young’s modulus of normal lymphocyte cells (0.471±0.081 kPa) was nearly four times higher than that of Jurkat cells (0.0964±0.0229 kPa) at the same loading rate. The stiffness of normal lymphocyte cells was (2.278±0.488) mN/m and that of Jurkat cells was (4.322±0.382) mN/m. The differences in mechanical properties of normal and cancerous cells were obvious that healthy and diseased states could be clearly distinguished. These results may be applied to the clinic disease diagnosis for distinguishing the normal cells from the cancer ones even when they show similar shapes.