FORM 2
THE PATENT ACT 1970
(39 of 1970)
&
THE PATENT (AMENDMENT) RULES, 2006
PROVISIONAL SPECIFICATION
(See section 10 and rule 13)
1. TITLE OF THE INVENTION
Single-Step Sample Processing Device
2. APPLICANT
(a) NAME : Span Diagnostics Ltd.
(b) NATIONALITY : IN
(c) ADDRESS : 173-B, New Industrial Estate, Udhna, Surat - 394210, India


PROVISIONAL The following specification describes the nature of this invention.
3. PREAMBLE TO THE DESCRIPTION

Field Of Invention:
The present invention relates to a single-step sample-processing device to improve testing in a rapid flow through test for detection of antibodies/antigens for infectious diseases in a given sample. This product will serve as an alternative to a multi-step existing flow through testing, having an additional advantage of being less time consuming than the other techniques and being more user friendly.
Background of the Invention:
Markets are flooded with various diagnostic test devices, which are available in different formats i.e. ELISA (Enzyme Linked Immuno Sorbent Assay), Fluorescent assay, immunodot based assay, Immunofilteration Assay and Lateral Flow Assays.
Ftow through assay is very well known format, which serves as a platform for diagnosis of infectious diseases. Various types of membrane based assays have been used and described in US patent S006464 to Chu et. al., US patent 4818677 to Hay-Kaufmann et. ah., US patent 4632901 to Valkris et. al., US patent 5185127 to Vonk et. al. and US patent 6284194 to Chu et. al.
Flow Through Assays for diagnosis of infectious diseases comes In different formats e.g. (a) device with test area to detect single analyte with or without control area (b) device with multiple test area to detect multiple analytes in one test with or without control artea. (in both cases analyte includes detection of antigen or antibody). To detect one has to apply sample, wash buffer and conjugate separately, in multi steps in the test device.
Despite being rapid Flow Through assay, they suffer from many disadvantages like:
• Multi-step testing procedure.
• Requires refrigeration temperature for storage.

the object of the present invention is to provide a single step sample-processing device, Which improves detection of antibodies/antigens In the test samples for Infectious diseases to obviate the drawbacks of the prior art.
This sample-processing device is more advantageous to multi-step application of sample, wash buffer and conjugate separately into the test device to detect antibody/ antigen in human serum and plasma. This device will provide single step improved testing, thereby minimizing testing time and is user friendly to apply sample, wash buffer and conjugate together, for rapid detection of infectious disease and show better performance in terms of sensitivity and specificity.
The invention will now be described with reference to the accompanied drawings.
Figure 1 shows cross-section of sample processing device according to this invention.
Figure 2 shows the exploded view of the sample-processing device.
Figure 3 shows the flow chart for testing.
Description of the Invention:
The present invention relates to a single step sample-processing device to improve testing in rapid flow testing for detection of antibodies/antigens in the human serum and plasma for infectious diseases.
Sample Processing Device as shown in figure 1, for carrying out the test comprising closed assembly with upper part called Unit base (1), and lower part called Unit cap (2) made preferably from flexible plastic although one may use wide variety of organic or inorganic material, natural or artificial or combination of any polymers including but not limited to the available part. Inside the assembly is the filter membrane (3) e.g. nitrocellulose stick on the unit cap. Filter membranes (3) for the Unit cap (2) may be nitrocellulose membranes, nitrocellulose mixed esters, nylon membranes, polysulfonyl

based membranes supported on suitable matrix such as polycarbonate Afters, and the like. Manufacturers of membranes (3) include Pall/Gelman, GE Life Science, MDI and Millipore. The said filter membrane (3) has pore size of about 0.4 micron to 0.7 micron, preferably of 0.6 micron. The residual area of the fitter membrane (3) i.e. nitrocellulose can be saturated or blocked with blocking agents to prevent non-specific binding which typically include either single or combination of any of the these compounds: proteins, synthetic polymers and surfactants. The other proteinaoaous-blocking reagent may be Casein and like. The other proteinaceous-blocking reagent may be selolaurate, TRTTON XrlOO™ (t-octylphenoxypolyethoxyethanol), sodium dodecylsulfate, n-octyl-D-glucopyranoside, NQNIDETTM (octylphenyl ethylene oxide), sodium dioxycholate etc. in concentration of 0.01 to 1.0 % (v/v). Blocking agents are applied in a buffer solution to the membrane, includes Tris (hydroxymethyl) aminomethane/HCI (Tris/HCI), Tris/citrate, Tris/maieate, Tris/glyclne, phosphate buffer, HEPES, and other biological buffers in the correct pH range. Another component of the sample-processing device is conjugate pad (4) as shown in figure l & 2. Conjugate pad (4) for the Unit cap (2) may be made up of glass fiber or cellulose ester. Manufacturers of membranes include Pall/Gelman, GE Life Science and MDI. The pad is preferably treated with surfactant to make it hydrophilic, which is known in the art. Known surfactants are but not limited to selolaurate, TRTTON X-1001" (t-octylphenoxypolyethoxyethanol), sodium dodecylsulfate, n-octyl-D-glucopyranoside, NONIDETTM (octylphenyl ethylene oxide), sodium dioxycholate and the like in concentration of 0.001 to 1.0 % (v/v). This treated or processed conjugate pad (4) are dipped in a colloidal gold conjugate of optimum optical density and dried in low humidity facility and lyophilized for better stability of the conjugate pad (4).
Sample Processing Buffer along with the filter membrane (3) will serve separation of the particulate matter from the sample under field conditions where centrifuge is not available, to perform this test. The Sample Processing Buffer is the isotonic buffer containing anticoagulant (0.001 to 0.002mg/mL), Phosphate buffer of molarity of 5m M to 50mM, detergent (2 to 4%), protein stabilizers (0.1 to 2%) and preservative (0.001 to 0.1%).

To achieve the one step procedure for performing the assay as shown in Figure 3, Sample processing buffer is added to Unit base (1) and sample (serum/plasma) is applied to it. The content is mixed and unit cap (2) is applied to unit base (1). The processed sample is allowed to pass through the filter membrane (3) and conjugate pad (4). No further additional step is required to get the results-
Advantages of the present Invention:
The present invention provides cost effective single-step sample processing device to improve testing in rapid flow through test for detection of antibodies/antigens in a given sample for infectious diseases.