T 1655/16 (Colored solutions PCR mixture/THERMO FISHER SCIENTIFIC BALTICS) 05-05-2022

Admission of the main request

1. The main request was originally filed as auxiliary request 2 with appellant I's statement of grounds of appeal. Claims 1 to 3 of the main request are directed to a method of preparing a reaction mixture for PCR assay which is identical to the method of claims 1 to 3 of auxiliary requests 5 and 6 at first instance, the latter request being that upheld by the opposition division. Claims 4 to 13 of the main request are directed to a solution set for PCR assay, wherein the feature in claim 4 "... the mixed solution comprises fluorescent agent and ... the absorbance peaks of any of said colorants do not overlap with the emission or excitation wavelength of said fluorescent agent" is the subject-matter of granted claim 13.

2. The patent proprietor maintained the granted claims as the main request throughout the whole proceedings at first instance and filed several auxiliary requests in response to the opponent's notice of opposition (auxiliary requests A to D) and the opposition division's preliminary opinion (auxiliary requests 1 to 4). At the oral proceedings at first instance, the opposition division admitted into the proceedings a new auxiliary request 4 to replace former auxiliary request 4 as well as new auxiliary requests 5 and 6 (the latter request being that upheld by the opposition division).

3. In all these auxiliary requests, amendments were introduced into claims 1 and 4. However, none of these auxiliary requests contained the subject-matter of granted claim 13. Although the amendments introduced into claim 4 of these auxiliary requests were mainly subject-matter taken from the description of the patent, claim 4 of some of these requests (auxiliary request D, former and new auxiliary request 4, and auxiliary request 5) was amended by introducing the subject-matter of granted claim 9, although slightly modified. Whilst the feature introduced into claim 4 of these auxiliary requests reads "... wherein the concentration of said colorants in respective reagent solutions corresponds to absorbances of the said solutions at their maximum absorption wavelength of 0.001-0.5 at 1 mm light path, when diluted to the desired PCR processing concentration", claim 9 as granted further refers to the preferred ranges "in particular 0.01 - 0.5, preferably 0.03 - 0.15".

4. The patent proprietor had thus ample opportunity to file claim requests during the first instance proceedings, including at the oral proceedings before the opposition division. The patent proprietor made use of these opportunities and filed several auxiliary requests, wherein amendments introduced into the independent claims were subject-matter taken from the description of the patent as well as from the granted claims. In view thereof, the board considers that the main request with the amendment introduced into claim 4 (subject-matter of granted claim 13) could and should have been filed at the first instance proceedings.

5. Appellant I argues that the filing of the main request in appeal proceedings was the first opportunity for the patent proprietor to react to the late filing of document (10) at the oral proceedings at first instance and the admission by the opposition division of this late filed document into the proceedings. The board does not agree therewith.

5.1 Document (10) was filed at the beginning of the oral proceedings at first instance during the discussion of whether the granted claims fulfilled the requirements of Article 54(3) EPC (cf. page 1, point 2.2, of the minutes of the oral proceedings at first instance; page 6, point 1, of the decision under appeal). Document (10) discloses the absorption maximum and extinction coefficients (epsilon) of the most commonly used quenchers, including Dabsyl (Table 6.2), a quencher referred to in document (1), the document cited under Article 54(3) EPC in the decision under appeal (cf. page 6, point 2, of the decision under appeal).

5.2 The opposition division stated that the disclosure of document (10) "was essential for the assessment of novelty over D1 in particular for a complete appreciation of the disclosure of Example 1 therein". At the same time, the document was considered to reflect the skilled person's general common knowledge in the field of fluorescent dyes and quenchers (cf. page 1, point 2.2, of the minutes of the oral proceedings at first instance; page 6, point 1.3, of the decision under appeal). The board agrees with this assessment. The nature of document (10) falls within the notion of common general knowledge as developed by the case law (cf. "Case Law of the Boards of Appeal of the EPO", 9th edition 2019, I.C.2.8.1, 77); the document provides only evidence of the skilled person's common general knowledge when reading the prior art, in particular Example 1 of document (1). The filing and content of document (10) did not raise any new facts or issues. Moreover, the patent proprietor was also given the opportunity to file new auxiliary requests once document (10) had already been admitted into the proceedings. As shown above, the patent proprietor had replaced auxiliary request 4 and filed new auxiliary requests 5 and 6.

5.3 In light of the course of events at first instance and the nature and disclosure of document (10), the board considers that the filing of the main request in appeal proceedings is an attempt to re-open the opposition proceedings, which is not in line with the purpose of an appeal proceedings as defined in the case law (cf. "Case Law", supra, V.A.1.1, 1133).

6. Thus, the board, in the exercise of its discretion (Article 25(2) RPBA 2020 and Article 12(4) RPBA 2007), decides not to admit the main request into the appeal proceedings.

Admission of new documentary evidence

7. Claim 1 of the main request is identical to claim 1 of auxiliary requests 5 and 6 filed at oral proceedings at first instance. The opposition division considered the feature introduced into claim 1, namely "wherein the third color is visually detected", to be "simple enough and also foreseeable in view of the teaching of the entire patent for the opponent not to be surprised or confronted with major difficulties" (cf. page 15, point 12.3, of the decision under appeal). The introduction of this feature was also considered to overcome the novelty objection over document (1) (cf. page 17, point 14.3, of the decision under appeal).

8. According to the minutes of the oral proceedings at first instance, the admission of auxiliary request 5 into the proceedings was discussed with the parties and, after a short break, the request was admitted into the proceedings (cf. page 2, point 5.1, of the minutes of the oral proceedings). The decision under appeal refers to both the opponent's reasons against the admission of this auxiliary request and those given by the opposition division for the admission of auxiliary request 5 into the proceedings (cf. page 15, points 12.2 and 12.3, of the decision under appeal).

9. According to the decision under appeal, the opponent referred only to the fact that auxiliary request 5 "was filed late and could have been provided earlier" (cf. page 15, point 12.2, of the decision under appeal). Neither in the minutes of these oral proceedings nor in the decision under appeal, there is any reference to the opponent arguing that the feature introduced into claim 1 of auxiliary request 5 resulted in a turn of events that required an interruption or postponement of the oral proceedings for the opponent to carry out a new search of prior art documents.

10. Appellant II's argument for the introduction of the new documentary evidence into the appeal proceedings based on the relevance of the feature introduced into claim 1 of auxiliary request 5 at first instance (main request in appeal), was put forward at a late stage of the appeal proceedings, namely in response to the board's provisional opinion not to admit this evidence into the appeal proceedings. In the statement of grounds of appeal, appellant II stated only that the new documents "are submitted on time as they are prima facie relevant and they are filed to demonstrate that the decision taken by the OD on inventive step was not correct. All these documents were therefore filed at the earliest possible time ... and therefore admissible". There was thus a reference only to the relevance of this new documentary evidence in the light of the opposition division's decision on Article 56 EPC.

11. The board agrees with appellant II that the relevance of the new documentary evidence must always be taken into account. However, in line with the case law, the relevance of new evidence cannot be the sole, decisive reason for a board to admit new evidence into the proceedings; the specific circumstances and particular course of events of the proceedings at first instance must also be taken into account (cf. "Case Law", supra, V.A.4.5.2, 1216). In the present case, several problem and solution approaches - based on the new documentary evidence - are formulated by appellant II as possible alternatives to the approach formulated by the opposition division in the decision under appeal and based on the combination of the closest prior art document (2) with document (3). Appellant II failed to provide any convincing reasons why the new documentary evidence and the alternative approaches based thereupon could not have been filed at an earlier stage of the proceedings at first instance.

12. The relevance of the feature introduced into claim 1 of auxiliary request 5 at first instance (main request in appeal) is derivable from the numerous references throughout the whole description of the patent, such as "one can tell directly by the color of the solution, whether it is the first reagent solution, the second reagent solution or the mixture of these" (cf. paragraph [0024]), "the inspection of the color can be made visually or by automatic optical means" (cf. paragraph [0026]), "the term "color" herein means any detectable spectral response (of a solution) to white light in the visual range ... which provides a colored visual appearance for the solution ... the colors are distinguishable, preferably by the naked eye ... capable of giving the solution a perceivable color" (cf. paragraphs [0050] to [0052], [0064], [0065], [0067] and [0093]), as well as by the examples of the patent, wherein visual colors are exemplified (cf. paragraph [0104] and [0120]).

13. Moreover, although the feature introduced into claim 1 of auxiliary request 5 at first instance (main request in appeal) was not present in any of the auxiliary requests filed in the written phase of the opposition proceedings, references were made throughout these proceedings to the "appearance" and the detection of a third color resulting from the mixing of first and second reagent solutions containing first and second colors (cf. page 3, point 2.1, and page 4, point 3.2, of the opposition division's preliminary opinion).

14. Thus, in view of the whole content of the patent and the course of events at first instance, the filing of auxiliary request 5 at oral proceedings at first instance with claim 1 amended by the introduction of the feature "the third color is detected visually" neither amounted to, nor can be seen as, a late turn of events requiring a search of new prior art documents and the interruption or postponement of these oral proceedings at first instance. Neither the former nor the latter were ever requested by the opponent at first instance. Therefore, the board considers that the first opportunity for the appellant II to react to the introduction of the feature "the third color is detected visually" was not in appeal proceedings but at an earlier stage of the proceedings at first instance.

15. Thus, the board, in the exercise of its discretion (Article 25(2) RPBA 2020 and Article 12(4) RPBA 2007), decided not to admit the new documentary evidence into the appeal proceedings.

Auxiliary request 1

Admission of the objections of lack of clarity and novelty

16. Claim 1 of auxiliary request 1 is identical to claim 1 of auxiliary requests 5 and 6 filed at the oral proceedings at first instance (the latter request being that upheld by the opposition division). The opposition division considered the subject-matter of this claim to be novel over document (1) (Article 54(3) EPC) (cf. page 17, point 14.3, of the decision under appeal; page 3, point 5.4, of the minutes of the oral proceedings at first instance). According to the minutes of the oral proceedings at first instance, after discussing the admissibility of auxiliary request 5 and the requirements of Article 123(2) EPC, the opponent did not raise any objection under Article 84 EPC and none was raised by the opposition division (cf. page 3, point 5.3, of the minutes of the oral proceedings at first instance). Since auxiliary request 6 was based on claims 1 to 3 and 15 of auxiliary request 5, auxiliary request 6 was considered to satisfy the requirements of Articles 123(2), 84 and 54 EPC without further discussion (cf. page 3, point 6.2, of the minutes of the oral proceedings at first instance). In line therewith, there is no reference to Article 84 EPC as regards auxiliary requests 5 and 6 in the decision under appeal.

17. In the statement of grounds of appeal, the sole decision of the opposition division contested by appellant II was the decision taken on Article 56 EPC. There was no reference to any other article of the EPC nor to document (1), the sole document cited under Article 54(3) EPC in the decision under appeal. Nor did appellant II refer to any of Articles 84 and 54 EPC in response to the board's communication informing the parties of the board's provisional opinion on the issues of the case. Appellant II only argued therein on the reasons for the admission of the new documentary evidence filed in the appeal proceedings.

18. It was only after informing the board of a change of representative that appellant II filed a submission with objections under Articles 84 and 54 EPC and new lines of attack against the requests on file. The introduction of these new objections and lines of attack was justified by appellant II by reference to the comments and amendments made by appellant I in the statement of grounds of appeal and concerning former auxiliary request 1. No other reasons were given by appellant II to explain the introduction of the new objections and lines of attack at that late stage of the appeal proceedings.

19. The introduction of the new objections under Articles 84 and 54 EPC and the new lines of attack are an amendment of appellant II's appeal case within the meaning of Article 13 RPBA. Thus, in line therewith, these new objections and lines of attack are not to be taken into account unless there are exceptional circumstances justified by cogent reasons.

20. In light of the course of events described above, the board fails to see any exceptional circumstances that may justify the admission of the new objections and lines of attack under Articles 84 and 54 EPC. Appellant I's comments and amendments referred to by appellant II were made at the earliest stage of the appeal proceedings and appellant II had ample opportunity to put forward the new objections and lines of attack at an earlier stage of these proceedings. The reasons given by appellant II are neither cogent nor justify the admission of the new objections and lines of attack at such late stage of the appeal proceedings.

21. The board also disagrees with appellant II's argument that the new objections and lines of attack are fundamental and based on a factual and evidential framework identical to that already present in the appeal proceedings. The framework of appeal proceedings is not only set out and defined by the decision under appeal alone but also by the reasons and evidence provided in the statement of grounds of appeal. As stated above, the purpose of appeal proceedings is not to re-open and continue the opposition proceedings but to review the decision of the opposition division (cf. "Case Law", supra, V.A.1.1, 1133). Appellant II's new objections and lines of attack under Articles 84 and 54 EPC are nothing more than an attempt to re-open the opposition proceedings and thus, not admissible.

22. Thus, the board, in the exercise of its discretion (Article 13 RPBA 2020), did not admit appellant II's new objections and lines of attack under Articles 84 and 54 EPC into the appeal proceedings.

Article 56 EPC

23. The closest prior art document (2) relates to the same technical field as the patent, namely PCR amplification of nucleic acids, and refers to several problems of the PCR method, in particular those due to the evaporation of the PCR solution and those arising from the application of PCR on a large scale (cf. page 3, third paragraph, to page 4, first paragraph). In this context, reference is made to the fact that specialised staff is required and that "the operator must ... prepare in advance all the reagents necessary for setting up the PCR reaction and aliquot into each of the wells of the plate the correct volume of amplification mixture, with all the risks deriving from the variability due to the operator" (cf. paragraph bridging pages 3 and 4).

24. As a solution to these problems, document (2) discloses the preparation of a master mix which is "mixed with an effective amount of an inert temperature controllable polymer" and which may comprise "all or part of the reagents required for the PCR amplification in the appropriate quantities" (cf. page 4, second and third paragraphs; claim 1). In one embodiment, the "master mix also comprises an effective amount of one or more inert dyes, preferably methylene blue or malachite green" (cf. page 6, fourth paragraph; claim 10). The teachings of document (2) are exemplified by the "reference dye (ROX)" (cf. page 9), known in the art as the inert fluorescent dye carboxyrhodamine.

25. Document (2) states that the amplification of a plurality of different target nucleotide sequences using a different dye for each target nucleic acid sequence "can be envisaged" (cf. paragraph bridging pages 6 and 7). However, there is no reference to a mixing of first and second reagent solutions with first and second colorants, respectively, and to the appearance of a third color - due to the mixing of said first and second colorants, which is different from those of the first and second colorants and which is detected visually, let alone a reference to the use of such a mixed solution as a reaction mixture for a PCR assay.

26. Starting therefrom, the objective technical problem is formulated as the provision of an alternative method for avoiding errors in the preparation and aliquoting of reaction mixtures for PCR amplification assay. There is no reference in document (2) to the detection of these errors. Consequently said detection cannot be part of the problem to be solved, otherwise hindsight knowledge of the patent would be implied.

The claimed method solves this problem and further allows the detection of errors, an effect that results from the proposed solution.

27. As stated above, document (2) refers to the presence of one or more inert dyes in the master mix (cf. page 6, fourth paragraph). An inert dye, such as the exemplified fluorescent dye ROX, is known in the field to be an internal passive reference which is not affected by PCR amplification and which allows for a normalisation of the PCR data. There is no hint or pointer in document (2) that could lead a skilled person to use first and second PCR reagent solutions with first and second colorants that, when mixing, result in a PCR reaction (master) mixture of a third color different from those of the first and second colorants and which is detected visually. Hindsight knowledge of the patent is required for a skilled person to envisage such a use from the disclosure of document (2) alone. Likewise, hindsight knowledge of the patent is required for combining the disclosure of document (2) with that of document (3).

28. Document (3) is not concerned with PCR amplification but with the isolation of nucleic acids, in particular plasmid DNA from E. coli and other organisms (cf. inter alia, abstract, paragraph [0003]). It is only after carrying out the disclosed purification method that the isolated pure plasmid DNA may be used, inter alia, in a PCR assay (cf. paragraphs [0005] and [0023]). Thus, it is arguable whether documents (2) and (3) are indeed in the same technical field so that a skilled person, when starting from the former, would be actually aware of the teachings of the latter document.

29. Moreover, whilst document (3) refers to the problems associated with inefficient sample handling related to pipetting (cf. paragraph [0011], penultimate sentence; paragraph [0012]), the disclosure of this document is engaged and deals with the lysis of E. coli and other bacterial organisms, in particular a modified mild alkaline lysis, which is an earlier step of the method for isolating plasmid DNA disclosed in document (3) (cf. inter alia, paragraphs [0006], [0007], [0031], [0047]; claims 1 and 8).

29.1 The contribution of document (3) to the art is the "addition of indicator dyes to the alkaline lysis based buffers [which] allows for easy and rapid visual monitoring of the resuspension, lysis, and neutralization steps. The ability to assess the process increases plasmid yield and also reduces errors" (cf. paragraph [0003]). As stated in document (3), an "improved visual recognition of lysis" or a "visual monitoring" of the modified mild alkaline lysis improves the "overall quality and yield" and "increases efficiency and significantly reduces the time required to process each sample" (cf. inter alia, paragraph [0011], last sentence; paragraphs [0012], [0023] and [0031]).

29.2 Although document (3) refers to indicator dyes that are "different in each of the buffers and may not undergo a response to pH", in the preferred embodiments "the color change is based on indicator dyes whose color is related to the pH of the solution" (cf. paragraph [0012]), and the preferred dyes "are ones that display different color ranges in the pH range suitable for plasmid DNA isolation" (cf. paragraph [0013]; claims 2 and 9).

29.3 Indeed, the colors of the indicator dyes present in the three buffers exemplifying the teachings of document (3) undergo a response to pH, namely buffer P1 (Resuspension buffer - with Phenol red, around neutral pH 7.0-8.0, red in color), buffer P2 (Lysis buffer - with Emerald Green, basic pH, blue-green in color) and buffer P3 (Neutralisation buffer - with Phenol Red, acidic pH, yellow in color). It is worth noting here that the same colorant is used in buffers P1 and P3 (cf. paragraphs [0013], [0016] to [0018], [0027], and [0033] to [0045]). The color changes reported in Example 1 of document (3) are based on these indicator dyes which are related to the pH and thus reflect the pH of the solutions after mixing. In Example 1, the addition of buffer P2 (basic pH, blue-green color) to buffer P1 (neutral pH, red color) results in a solution of "a deeper red in color" (cf. paragraphs [0051] and [0052]), and the addition of buffer P3 (acidic pH, yellow color) to the latter solution results in a final solution that "becomes yellow" (cf. paragraph [0053]).

29.4 It is questionable whether a skilled person would have considered the preferred strategies described and exemplified in document (3) for visual monitoring of a modified mild alkaline lysis of E. coli host cells based on pH dependent colorants for solving the objective technical problem as formulated above and arriving thereby at the claimed subject matter. The board agrees with the decision of the opposition division and considers that this question must be answered in the negative (cf. page 20, point 16.4, of the decision under appeal).

30. Therefore, the skilled person would not have combined the teaching of documents (2) and (3) and, even if they were combined, hindsight knowledge of the patent would have been required for a skilled person to arrive at the method claimed in auxiliary request 1.

31. Thus, auxiliary request 1 fulfils the requirements of Article 56 EPC.